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  • 1985-1989  (131)
  • 1955-1959  (7)
  • 1940-1944
  • Agrobacterium
  • gene expression
  • 1
    ISSN: 1432-2048
    Keywords: Agrobacterium ; Auxin and thiophene in roots ; Root culture ; Root morphology ; Tagetes ; Thiophene ; Transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Roots of marigold (Tagetes patula L.) accumulate thiophenes, heterocyclic sulfurous compounds with strong biocidal activity. In detached roots cultured in vitro, the thiophene content was 5 μmol·(g fresh weight)-1 which is 25-times higher than in roots attached to the plant. In roots derived from tissues transformed by Agrobacterium tumefaciens and A. rhizogenes, the morphology and thiophene content varied with the bacterial strain used. Transformation stimulated the elongation of the root tips and the formation of lateral roots but lowered the thiophene level to 20–50% relative to the concentration in untransformed detached roots. A negative correlation was found between the number of laterals in a root system and the thiophene content. Extensive branching and a decrease in thiophene accumulation was evoked in untransformed roots by indole-3-acetic acid (1–10 μmol·l-1) added to the medium. Within the roots, the highest thiophene concentrations were found in the tips. The results indicate that auxin directly or indirectly plays a role in the regulation of the thiophene level in root tips.
    Type of Medium: Electronic Resource
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  • 2
    Electronic Resource
    Electronic Resource
    Springer
    Cellular and molecular life sciences 45 (1989), S. 526-535 
    ISSN: 1420-9071
    Keywords: Interferon ; cytokines ; interleukins ; gene expression ; transcription ; autoimmune ; disease
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Specific interferon genes are transcribed at low levels in the spleen, liver, and peripheral blood leukocytes of normal individuals in the apparent absence of virus infection while other interferon genes remain unexpressed in the same tissues. In contrast, the genes of cytokines such as IL-1, IL-6 and TNF are expressed at relatively high levels in the organs of normal individuals. The level of expression of the IL-1, IL-6 and TNF genes is markedly reduced in the livers of patients with autoimmune liver disease compared to the level of expression in the liver of normal individuals, whereas the expression of interferon genes is similar in both normal and diseased liver, suggesting that a defect in the expression of specific cytokines is associated with severe liver disease.
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  • 3
    ISSN: 1432-203X
    Keywords: Agrobacterium ; Somatic Embryogenesis ; Electroporation ; β-Glucuronidase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Daucus carota hypocotyl sections were transformed withAgrobacterium tumefaciens LBA4404 containing CaMV 35S promoter, β-glucuronidase coding sequence and the nopaline synthase (Nos) poly adenylation sequences in Bin 19. Sliced sterile seedling hypocotyl segments were preincubated for 2 days, co-cultivated withAgrobacterium for an additional 2 days, and then transferred to medium containing 100ug/ml of kanamycin and 400ug/ml carbenicillin. In 6 weeks kanamycin resistant calli were obtained in 5.8% of the explants from one variety. Calli were subcultured on solid medium, and in 4 weeks introduced into suspension culture. NPTII and Southern blot analysis confirmed that three selected lines were transformed with 1–3 copies of the GUSII construction. GUS activity in transformants was 5 to 250 fold over background.
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  • 4
    ISSN: 1572-994X
    Keywords: BAL31 nuclease ; CAT assay ; gene expression ; HTLV-I ; R region ; 5′ untranslated sequence
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract We previously demonstrated the activity of the R fragment in the long terminal repeat of human T-cell leukemia virus type I for elevation of the level of gene expression. In this study, the fragment was deleted with BAL31 nuclease to determine its functional domain. Series of the shortened R fragments were linked to the simian virus 40 promoter unit, which regulated expression of a reporter gene. Examination with the R fragments deleted from the 5′ and 3′ ends showed that borders of the functional domain were mapped within nucleotide positions 458 to 473 for the 5′ end and nucleotide positions 559 to 594 for the 3′ end, respectively. Thus we conclude that a 136-base-pair fragment corresponding to the second half of the R region was sufficient to allow elevation of the level of gene expression.
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  • 5
    Electronic Resource
    Electronic Resource
    Springer
    Bulletin of experimental biology and medicine 108 (1989), S. 999-1001 
    ISSN: 1573-8221
    Keywords: brain ; gene expression ; mRNA ; transplantable hepatomas
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Type of Medium: Electronic Resource
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  • 6
    ISSN: 1573-4927
    Keywords: mouse ; salivary protein ; lacrimal protein ; gene expression ; Spt locus ; multigene family
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract The murine submandibular gland (SMG) produces a novel class of highly acidic salivary proteins encoded by one or more highly abundant mRNA transcripts. In inbred mice, these transcripts are encoded by members of a multigene family comprising approximately 8–12 homologues. Most, and probably all, of these homologues are clustered at a new locus near belted (bt) on chromosome 15, which we designateSpt (salivary protein). Although physically closely linked,Spt genes differ in their patterns of expression both in strains of mice and in their tissues. One gene,Spt-1, is expressed at high levels in the SMG of all inbred strains examined. This gene is also expressed at significant levels in the lacrimal gland. A second gene,Spt-2, appears to be present as a single copy in some strains and as two copies in others. This gene is expressed at high levels only in the SMG of those strains carrying two copies, andSpt-2 mRNA is not detectable in the SMG of strains carrying only one copy. In contrast toSpt-1, theSpt-2 gene is not expressed at detectable levels in the lacrimal gland.
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  • 7
    ISSN: 1573-4927
    Keywords: mouse ; salivary protein ; lacrimal protein ; gene expression ; Spt locus ; multigene family
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract The murine submandibular gland (SMG) produces a novel class of highly acidic salivary proteins encoded by one or more highly abundant mRNA transcripts. In inbred mice, these transcripts are encoded by members of a multigene family comprising approximately 8–12 homologues. Most, and probably all, of these homologues are clustered at a new locus near belted (bt) on chromosome 15, which we designateSpt (salivary protein). Although physically closely linked,Spt genes differ in their patterns of expression both in strains of mice and in their tissues. One gene,Spt-1, is expressed at high levels in the SMG of all inbred strains examined. This gene is also expressed at significant levels in the lacrimal gland. A second gene,Spt-2, appears to be present as a single copy in some strains and as two copies in others. This gene is expressed at high levels only in the SMG of those strains carrying two copies, andSpt-2 mRNA is not detectable in the SMG of strains carrying only one copy. In contrast toSpt-1, theSpt-2 gene is not expressed at detectable levels in the lacrimal gland.
    Type of Medium: Electronic Resource
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  • 8
    Electronic Resource
    Electronic Resource
    Springer
    Cellular and molecular neurobiology 9 (1989), S. 281-294 
    ISSN: 1573-6830
    Keywords: cholecystokinin ; mammillary region ; development ; gene expression ; hybridization histochemistry
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary 1. The ontogeny and adult distribution of neurons containing cholecystokinin (CCK) mRNA in the premammillary and mammillary nuclei and supramammillary region of the rat brain were studied using hybridization histochemistry. 2. The earliest detection of CCK mRNA in the mammillary region was on E14, followed by a marked increase in transcript levels during the next 4 days, a time during which neurons in this region still divide. During the first 2 weeks of life, few changes in the levels of CCK transcripts were seen, and an adult-like pattern of expression was seen on the twenty-first day of life. 3. Low levels of transcripts were present in numerous neurons located in all divisions of the medial nucleus and in the posterior nucleus known to project ipsilaterally to the anteroventral and anteromedial thalamic nuclei. In contrast, none of the neurons in the lateral nucleus (projecting bilaterally to the anterodorsal thalamic nucleus) had detectable transcripts. 4. Many neurons in the supramammillary nucleus had low, moderate, or high levels of transcripts. Some nearby nuclei (such as the dorsal premammillary nucleus) had smaller numbers of neurons with low levels of CCK mRNA, whereas others (such as the ventral premammillary nucleus) had none.
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  • 9
    ISSN: 1573-0603
    Keywords: Agrobacterium ; gene transfer ; somatic embryos ; walnut ; β-glucuronidase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Somatic embryos have been successfully used as a target tissue for transformation and regeneration of transgenic walnut plants. Walnut somatic embryos, initiated originally from developing zygotic embryos, proliferate numerous secondary embryos from single cells in the epidermal layer. These single cells in intact somatic embryos are susceptible to transformation by genetically engineeredAgrobacterium tumefaciens and provide a means to regenerate nonchimeric transgenic plants. This gene transfer system has been made more efficient using, a) vector plasmids containing two marker genes encoding β-glucuronidase (GUS) and aminoglycoside phosphotransferase (APH(3′)II) and B) a more virulent strain ofAgrobacterium. This system should be applicable to any crop that undergoes repetitive embryogenesis from singleAgrobacterium-susceptible cells.
    Type of Medium: Electronic Resource
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  • 10
    ISSN: 1573-5028
    Keywords: Agrobacterium ; auxins ; roots ; oncogenes ; grapevine ; iaa genes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A new type of root-inducing iaa gene set was cloned from the Ti plasmid of the biotype III Agrobacterium tumefaciens strain Tm-4. These iaa genes are characterized by a very low DNA homology with the well-characterized iaa gene set, iaaM and iaaH, of the “common DNA” region of the biotype I strain Ach5 and by a low root-inducing activity. The biological activities of both iaa gene sets were compared by transferring each into a disarmed Ti vector and by testing the resulting strains on Nicotiana rustica leaf discs, decapitated Datura stramonium stems, tomato plants and Kalanchoë daigremontiana. Tm-4 iaa genes have a reproducibly weaker root-inducing ability on Nicotiana rustica, induce very little tumour growth on decapitated Datura plants or on tomato plants and do not induce roots on Kalanchoë daigremontiana. The Tm-4 iaa region was mapped by λ:: Tn5 transposon mutagenesis and tested on Nicotiana rustica. These tests combined with complementation experiments map the iaa genes to a 4.5-kb region. The Tm-4 iaa genes were able to complement the corresponding Ach5 iaa genes on Nicotiana rustica, indicating that the differences between these genes are quantitative rather than qualitative. Complementation experiments on Kalanchoë showed the iaaM gene of Tm-4 responsible for the overall weak auxin activity of the intact iaa set. In view of the observed structural and functional differences we propose to call the Tm-4 iaa genes TB-iaaM and TB-iaaH and the Ach5 iaa genes A-iaaM and A-iaaH.
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  • 11
    Electronic Resource
    Electronic Resource
    Springer
    Plant molecular biology 12 (1989), S. 295-299 
    ISSN: 1573-5028
    Keywords: gene expression ; multiple transcripts ; phytochrome ; Pisum sativum ; red/far-red reversible effect
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The reversible effect of red and far-red light on the level of two RNA transcripts of the single-copy phytochrome gene in pea was investigated using the primer extension assay. In dark-grown seedlings, a brief irradiation with red light markedly reduced the level of one of the phytochrome transcripts, RNA1. This red light effect was reversed by subsequent irradiation with far-red light. The other phytochrome transcript, RNA2, was only slightly influenced by light. In light-grown seedlings, a brief irradiation with far-red light increased the content of both RNA1 and RNA2 when the seedlings were transferred from light to darkness. This far-red light effect was reversible by subsequent red light irradiation.
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  • 12
    Electronic Resource
    Electronic Resource
    Springer
    Plant molecular biology 13 (1989), S. 327-336 
    ISSN: 1573-5028
    Keywords: Agrobacterium ; plant cell transformation ; plant tumor induction ; plant vector systems
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 13
    ISSN: 1573-5028
    Keywords: antisense RNA ; β-glucuronidase ; tobacco ; transgenic plants ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Antisense RNA was used to specifically inhibit the expression of a GUS gene introduced in a transgenic plant. A tobacco transformant containing a single intact copy of the GUS gene and showing relatively high constitutive levels of GUS activity (GUS+) was re-transformed with an Agrobacterium Ti-derived binary vector containing an antisense version of this reporter gene. The sense and antisense GUS genes were each under the regulation of the CaMV 35S promoter. Re-transformed plants contained 1–5 copies of the antisense construct and all showed a greater than 90% reduction in GUS activity relative to the original GUS+ plant. This reduction in GUS activity correlated closely with the levels of GUS enzyme and steady state GUS mRNA observed in these plants. The relatively low levels of sense and antisense GUS transcripts found in the re-transformed plants may indicate a rapid degradation of the RNA:RNA duplex in the cell.
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  • 14
    ISSN: 1573-4919
    Keywords: complement ; factor B ; gene expression ; interferon-ψ ; interleukin-1
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The serine protease complement factor B (Bf), an acute phase plasma protein, is a component of the alternative pathway of complement activation. Previous studies revealed that several cytokines including IFN-γ and IL-1 are involved in mediating acute phase Bf expression. To determine the molecular details of Bf expression we isolated, sequenced and characterized the 5′ flanking regions of the human and murine Bf genes. In both species the Bf transcriptional start site in liver was located 〈400 by 3′ to the polyadenylation site of the upstream C2 gene. This upstream intergenic region contained 〉65% nucleotide homology between species. Within this region, an IRS and three heat shock consensus elements were found in the murine sequence in an identical position to that of the human. To examine the functional details of Bf expression, a series of mouse and human Bf promoter - chloramphenicol acetyltransferase (CAT) chimeric gene constructs were transfected into mouse L or human HepG2 cells. Analysis of expression of these fusion gene constructs revealed that 1) cis-acting DNA sequences identified, at least in part, in the 3′ untranslated region of the C2 gene (within the 400 by upstream of the Bf cap site) mediate responsiveness to IL-1 and IFN-γ, 2) the responsiveness to each mediator appears to be conferred by separate upstream regions similar in position and homologous in man and mouse, and 3) the IL-1 responsive region in both species appears to have the characteristics of an enhancer element. The results of this analysis suggest a selective pressure to conserve the intergenic sequence between C2 and Bf genes and that further studies of these sequences will be useful in elucidating mechanisms controlling the acute phase response.
    Type of Medium: Electronic Resource
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  • 15
    Electronic Resource
    Electronic Resource
    Springer
    Bioscience reports 9 (1989), S. 157-188 
    ISSN: 1573-4935
    Keywords: matrix hypothesis ; morphogenesis ; protodynamics ; growth control ; DNA ; genome organisation ; gene expression ; gene regulation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract A theoretical concept is proposed, in order to explain some enigmatic aspects of cellular and molecular biology of eukaryotic organisms. Among these are the C-value paradox of DNA redundancy, the correlation of DNA content and cell size, the disruption of genes at DNA level, the “Chromosome field” data of Lima de Faria (Hereditas 93∶1, 1980), the “quantal mitosis” proposition of Holtzeret al. (Curr. Top. Dev. Biol. 7∶229 1972), the inheritance of morphological patterns, the relations of DNA and chromosome organisation to cellular structure and function, the molecular basis of speciation, etc. The basic proposition of the “Unified Matrix Hypothesis” is that the nuclear DNA has a direct morphogenic function, in addition to its coding function in protein synthesis. This additional genetic information is thought to be largely contained in the non-protein coding transcribed DNA, and in the untranscribed part of the genome.
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  • 16
    ISSN: 1573-5028
    Keywords: cadmium ; Datura innoxia ; gene expression ; heat shock ; tolerance
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The effect of Cd on gene expression in suspension cultures of twoDatura innoxia cell lines with differing Cd tolerance was studied.In vivo labeling experiments using [3H] leucine showed that Cd induced the synthesis of a similar range of proteins in both cell lines at a concentration which will kill the sensitive but not the tolerant cells. Corresponding changes in levels of translatable mRNA were also observed. The induction of the synthesis of proteins by Cd was transient since Cd-tolerant cells growing continuously in 250 μM CdCl2 contained a similar set ofin vitro translation products to cells growing in the absence of Cd. Although Cd had a similar effect on gene expression in both cell lines, Cd-tolerant cells possess two abundant mRNAs which are constitutively produced. These mRNAs encode proteins of low molecular weight (about 11 kDa) and are either absent or present at a low level in Cd-sensitive cells. The functions of these proteins are not known but they may be involved in the tolerance mechanism. Two-dimensional gel electrophoresis ofin vitro translation products showed that many of the Cd-induced proteins are also induced by heat shock. A 42°C heat shock resulted in agreater range and more intense induction of translatable mRNAs than 4 h exposure to 250 μM CdCl2. However a subset of mRNAs were induced specifically by Cd while other mRNAs were heat shock-specific. There was no difference in the ability of the two cell lines to tolerate heat shock. This was also reflected by the same pattern of major proteins induced by heat shock in the two cell lines.
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  • 17
    Electronic Resource
    Electronic Resource
    Springer
    Plant molecular biology 12 (1989), S. 667-672 
    ISSN: 1573-5028
    Keywords: ferredoxin ; gene expression ; heterocyst ; Anabaena 7120 ; Escherichia coli
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The genes for ferredoxin from heterocysts (fdx H) and vegetative cells (pet F) of Anabaena sp. strain 7120 were subcloned into plasmid pUC 18/19. Both genes were expressed in Escherichia coli at high levels (≈10% of total protein). Pet F could be expressed from its own promoter. The ferredoxins were correctly assembled to the holoprotein. Heterocyst ferredoxin was purified from E. coli extracts on a large scale. Its biochemical and biophysical properties were identical to those of the authentic ferredoxin, isolated from Anabaena heterocysts.
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  • 18
    ISSN: 1573-5028
    Keywords: agroinfection ; Agrobacterium ; strains ; maize streak virus ; Gramineae ; maize
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Parameters affecting the efficiency of agroinfection of maize streak virus (MSV) in maize have been determined. Monomeric units, cloned at a number of sites in the MSV genome were not infectious but multimeric units containing partial duplications were equally as infectious as complete tandem dimeric clones. Inoculation of tandem dimeric units conjugated into different strains of Agrobacterium showed that both A. tumefaciens and A. rhizogenes were able to transfer DNA to maize and this ability was Ti (or Ri) plasmid-specific. Nopaline strains of A. tumefaciens and both agropine and mannopine A. rhizogenes strains efficiently transferred MSV DNA to maize. A number of strains were capable of MSV DNA transfer to other members of the Gramineae, providing information which may be essential for Agrobacterium-mediated transformation of monocotyledonous plants.
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  • 19
    ISSN: 1573-5028
    Keywords: acyl carrier protein ; fatty acid synthesis ; gene expression ; gene fusion ; protein A ; spinach
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A synthetic gene encoding spinach acyl carrier protein I (ACP-I) was fused to a gene encoding the Fc-binding portion of staphylococcal protein A. This gene fusion, under the control of the λPR promoter, was expressed at high levels in Escherichia coli producing a 42 kDa fusion protein. This fusion protein was phosphopantethenylated in E. coli. In vitro the ACP portion of the fusion protein was able to participate in acyl ACP synthetase reactions, plant malonyl-CoA:ACP transacylase (MCT) reactions, and plant fatty acid synthetase (FAS) reactions. Inhibitory effects of high ACP concentrations on in vitro plant FAS were observed with the unfused ACP-1 but not with the fusion protein. As with unfused ACP-I, the fusion protein was a poor substrate for E. coli FAS reactions. When injected into rabbits, the fusion protein was also able to generate antiserum to spinach ACP-I.
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  • 20
    ISSN: 1573-5028
    Keywords: cDNA sequence ; gene expression ; glutamine synthetase ; legume-Rhizobium symbiosis ; nitrogen metabolism ; Phaseolus vulgaris L.
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We report the sequence of an essentially full-length glutamine synthetase (GS) cDNA clone (pcGS-γ1) isolated from a root nodule library ofPhaseolus vulgaris L. The polypeptide encoded by this cDNA has been producedin vitro by transcription/translation and shown to co-migrate on two-dimensional gels with the previously identified major cytosolic GS polypeptide (γ) of nodules. Two previously identified GS cDNA clones, pR-2 and pR-1 (see Gebhardtet al., EMBO J 5: 1429–1435, 1986) have similarly been shown to encode the α and β cytosolic GS polypeptides respectively. An RNase protection technique has been used to analyse specifically and quantitatively the abundance of mRNA related to these three GS cDNAs and to the cDNA (pcGS-δ1) encoding the chloroplast-located GS, during nodulation. Differences in the abundances of these mRNAs at different times suggest that they are not coordinately regulated. Moreover, using this technique mRNA specifically related to pcGS-γ1 was found at high levels in nodules but not in roots or leaves. Surprisingly the expression of this gene is not nodule-specific as previously suggested, as its mRNA was also detected, but at lower levels, in stems, petioles and in green cotyledons. By comparison, mRNA related to a leghaemoglobin gene was detected only in nodules. Comparisons of the relative abundances of the pcGS-γ1 mRNA and the γ polypeptide in different organs and at different stages during nodulation, suggest that the appearance of the γ polypeptide is largely under transcriptional control.
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  • 21
    ISSN: 1573-5028
    Keywords: Agrobacterium ; hairy roots ; T-DNA ; geotropism
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Single and multiple infections of carrot discs were carried out with Agrobacterium strains harbouring different segments of pRi1855 TL-DNA cloned in the binary vector Bin 19 and with a strain carrying the TR-DNA from the same Ri plasmid. Roots induced by the various co-inoculations were cultured and their growth patterns were followed. Abundant roots could be induced by TL-DNA rol genes A, B and C as a single insert (rolA+B+C) and by rolB alone provided an extended segment beyond its 5′ noncoding region was included in the construction. A depression of rooting capability was caused by the inclusion of rolC together with rolB (rolB+C). In all cases co-inoculation with the Agrobacterium carrying TR-DNA-borne auxin genes was necessary for root induction since none of the rol constructions was in itself capable of eliciting any response; an exceeding majority of these roots were however shown to contain rol genes but no TR-DNA. Rooting was also elicited if rol constructions were co-inoculated with a strain carrying TL-DNA genes 13 and 14 (ORF13+14) instead of the TR-DNA strain. These roots were shown to contain both rol genes and ORF13+14. Striking differences in growth properties were shown by roots containing different complements of TL-DNA genes. Typical hairy root traits, high growth rate, branching and, most noticeably, absence of geotropism, were shown by roots containing rolB alone, while roots with rolA+B+C were geotropic as normal carrot roots. Hairy root traits were conferred to rolA+B+C roots by the concomitant presence of ORF13+14 and by the addition of auxin to the culture medium. A model is presented which attempts to rationalize the growth patterns by assigning interplaying roles to the various TL-DNA genes involved.
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  • 22
    ISSN: 1573-5028
    Keywords: differential cDNA screening ; gene expression ; hybrid-select in vitro translation ; nuclear run-off transcription ; RNA blot hybridization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A cDNA library from cultured parsley (Petroselinum crispum) cells was differentially screened using labeled run-off transcripts derived from nucleic of elicitor-treated and untreated cells. This resulted in the isolation of 18 independent cDNA families representing putative defense-related genes. All genes are rapidly and transiently activated after elicitor application, but the time courses of transcriptional activity exhibit considerable variations, indicating differences in the mechanisms of gene regulation.
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  • 23
    ISSN: 1573-5028
    Keywords: deformation factor ; gene expression ; pea ; Rhizobium leguminosarum ; root hairs
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The mRNA population in pea root hairs was characterized by means of in vitro translation of total root hair RNA followed by 2-dimensional gel electrophoresis of the translation products. Root hairs contain several mRNAs not detectable in total RNA preparations from roots. Most of these root hair-specific mRNAs occur in elongating root hairs at higher levels than in mature root hairs. The expression of some genes in pea root hairs is typically affected by inoculation with Rhizobium leguminosarum. One gene, encoding RH-42, is specifically induced while the expression of another gene, encoding RH-44, is markedly enhanced. Using R. leguminosarum mutants it was shown that the nodC gene is required for the induction and enhancement of expression of the RH-42 and RH-44 genes, respectively, while the Rhizobium chromosomal gene pss1, involved in exopolysaccharide synthesis, is not essential. After induction of the nod genes with apigenin the bacteria excrete into the culture medium a factor that causes root hair deformation. This deformation factor stimulates the expression of the RH-44 gene but does not induce the expression of the gene encoding RH-42.
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  • 24
    ISSN: 1573-5028
    Keywords: C4 photosynthesis ; gene structure ; gene expression ; genetic variation ; silent substitution ; Zea mays
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have determined the structure of the maize (Zea mays L. subsp.mays line B73) nuclear gene encoding the phosphoenolpyruvate (PEP) carboxylase isozyme involved in C4 photosynthesis. The gene is 5.3 kb long and has ten exons that range in size from 85 to 999 bp. The nine introns vary from 97 to 872 bp. The sequence of 663 bp of 5′-flanking and 205 bp of 3′-flanking DNA is reported along with the entire gene sequence. Several short repetitive sequences were found in the 5′-flanking DNA that have characteristics similar to elements important in the light regulation of pea genes encoding the small subunit of ribulose 1,5-bisphosphate carboxylase. In addition, some 5′-flanking sequence similarities were found in a comparison with other light-regulated genes from maize and wheat. The level of DNA sequence variation among different PEP carboxylase alleles is similar to the allelic variation observed for several other maize nuclear genes. The data suggest modern maize variaties have retained much of the genetic variation present in their ancestral forms. Finally, accumulation of transcripts encoding the PEP carboxylase isozyme involved in C4 photosynthesis is quite high in several structures besides leaves, including inner leaf sheaths, tassels and husks. This indicates that expression of this gene is not leaf-specific and may not necessarily be coupled to the development of Kranz anatomy.
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  • 25
    Electronic Resource
    Electronic Resource
    Springer
    Plant molecular biology 12 (1989), S. 673-682 
    ISSN: 1573-5028
    Keywords: amylase inhibitor ; barley ; cDNA sequence ; gene expression ; hormonal regulation ; protease inhibitor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have cloned and sequenced a full-length cDNA from barley (Hordeum vulgare L.) seeds encoding the bifunctional α-amylase/subtilisin inhibitor (BASI). The nucleotide sequence predicts an open reading frame coding for a protein of 203 amino acids. The first 22 amino acids exhibit the sequence characteristic of a signal peptide, as found in several other plant protease inhibitors. Northern blot hybridization experiments indicate that BASI mRNA accumulation is strictly tissue-specific and is developmentally programmed. BASI mRNA transcripts were only identified in 1) developing starchy endosperm tissue from 14 days after flowering and 2) aleurone tissue of germinating seeds. In this latter tissue, BASI mRNA accumulation is enhanced by abscisic acid and abolished by gibberellic acid. Expression of BASI mRNA was also studied in the lys 3a high-lysine barley mutants Risø No. 1508 and Piggy. These high-lysine barleys show 2–4-fold higher levels as well as prolonged accumulation of BASI mRNA compared to the normal motherline Bomi. This correlates with the increased deposition of BASI protein in lys 3a barley mutants. Genomic blot analysis of barley DNA suggests that there are one or two BASI structural genes per haploid genome. Possible roles of BASI as part of a defence mechanism against precocious germination and pathogens are discussed.
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  • 26
    ISSN: 1573-5028
    Keywords: cDNA ; gene expression ; hydroxypyruvate reductase ; light regulation ; peroxisomal enzyme
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A full-length cDNA encoding NADH-dependent hydroxypyruvate reductase (HPR), a photorespiratory enzyme localized in leaf peroxisomes, was isolated from a λgt11 cDNA library made by reverse transcription of poly(A)+ RNA from cucumber cotyledons. In vitro transcription and translation of this clone yielded a major polypeptide which was identical in size, 43 kDA, to the product of in vitro translation of cotyledonary poly(A)+ RNA and subsequent immunoprecipitation with HPR antiserum. Escherichia coli cultures transformed with a plasmid construct containing the cDNA insert were induced to express HPR enzyme activity. RNA blot analysis showed that HPR transcript levels rise significantly in the first eight days of light-grown seedling development. This closely resembles the pattern seen for HPR-specific translatable mRNA. DNA blot analysis indicated that a single HPR gene is likely present per haploid genome. Nucleotide sequence analysis revealed an open reading frame of 1146 bases which encodes a polypeptide with a calculated molecular weight of 41.7 kDa. The derived amino acid sequence from this open reading frame is 26% identical and 50% similar to the amino acid sequence of the E. coli enzyme phosphoglycerate dehydrogenase, which catalyzes a similar reaction and functions in a related pathway. Statistical analyses show that this similarity is significant (z〉10). The derived amino acid sequence for HPR also contains the characteristics of an NAD-binding domain.
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  • 27
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    Plant molecular biology 13 (1989), S. 365-373 
    ISSN: 1573-5028
    Keywords: Agrobacterium ; binary vectors ; bleomycin ; phleomycin ; selective markers ; tobacco
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Tobacco cells are sensitive to bleomycin and phleomycin. The Tn5 and the Streptoalloteichus hindustanus (Sh) bleomycin resistance (‘Ble’) genes conferring resistance to these antibiotics have each been inserted into two plant expression vectors. They are flanked by the nopaline synthase (nos) or the cauliflower mosaic virus (CaMV) 35S promoters on one side, and by the nos polyadenylation signal on the other. These four chimaeric genes were introduced into the binary transformation vector pGA 492, which were thereafter mobilized into Agrobacterium tumefaciens strain LBA 4404. The resulting strains were used to transform Nicotiana tabacum cv. Xanthi nc using the leaf disc transformation procedure. In all cases, phleomycin- and bleomycin-resistant tobacco plants were regenerated from transformed cells under selective conditions; however the highest frequency of rooted plants was obtained when transformation was carried out with the ‘Sh Ble’ gene under the control of the 35S promoter. Phleomycin resistance was stably transmitted to sexual offspring as a dominant nuclear trait as confirmed by Southern blotting.
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  • 28
    ISSN: 1617-4623
    Keywords: Agrobacterium ; Hairy root ; T-DNA ; Glucuronidase gene
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Root differentiation could be elicited on carrot discs by transformation with the agropine Ri plasmid rolB gene cloned in the binary vector Bin19, provided two conditions were met. Firstly, an adequate auxin supply had to be provided. This was achieved by co-inoculation with a strain carrying only the auxin synthetic genes of the TR-DNA. Most of the resulting roots were then shown to harbour only rolB and no aux genes. Secondly, an extended non-coding region (∼1200 bp) at the 5′ end of rolB had to be included in the construction. A shorter (∼300 bp) 5′ region, including TATA and CCAAT boxes, was not sufficient to trigger root differentiation. Both the extended (B1185) and reduced (B310) 5′ regions of rolB were then cloned upstream of the β-glucuronidase (GUS) reporter gene and infections carried out both on the apical and on the basal side of carrot discs. Strong expression of GUS, visualized histochemically as an intense blue colouring of transformed cells was observed with B1185-GUS constructions on the apical side of the discs. Only occasionally could coloured cells be observed on the basal side of the discs with B1185-GUS and on both apical and basal sides with B310-GUS constructions. Strong GUS expression was, on the contrary, achieved on cells of both auxin-rich (apical) and auxin-depleted (basal) sides of the discs with the strong constitutive viral promoter, CaMV35S. These results indicate the presence of an upstream regulatory region which confers polar expression to the rolB gene and suggest a role for auxin in its activation.
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  • 29
    ISSN: 1615-6102
    Keywords: In vitro shoots ; Pisum sativum ; Meristematic cells ; Shoot apices ; Transformation ; Agrobacterium
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Different tissues in cultured pea shoots were inoculated withAgrobacterium tumefaciens wild types C 58 and ACH 5 andA. rhizogenes wild type 9402. The C 58 and 9402 bacteria induced the formation of tumours and hairy roots respectively while the ACH 5 was inactive. The younger the tissue the more rapidly it responded to the active bacteria. The shoot apex was the most reactive organ and developed into a tumour, theA. rhizogenes tumours subsequently giving rise to transformed hairy roots. Histological examination showed that transformed cells (including those within the apical dome) initially became highly vacuolate before dividing rapidly to form a tumour. These changes were accompanied by cell division in surrounding tissues.
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  • 30
    ISSN: 1617-4623
    Keywords: Agrobacterium ; IS elements ; Ti plasmid ; T-region ; Bacterial evolution
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A large number of Agrobacterium tumefaciens strains of the biotype III group carry two to ten copies of two related IS elements, IS866 and IS867. A study of the distribution and localization of these elements in 54 strains showed that one IS866 and two IS867 copies are always found at characteristic sites on the octopine/cucumopine and vitopine Ti plasmids, whereas varying amounts of IS866 and IS867 copies occur at different positions on the chromosome. By comparison of the IS patterns, an evolutionary tree could be deduced which shows the phylogenetic relationships between 23 different types of Agrobacterium strains. The structures of the T-regions of the different strains were also compared. Within the octopine/cucumopine group, eight T-region patterns could be defined. These patterns were found to be correlated with the chromosomal IS patterns. This strongly suggests that the IS866 and IS867 containing Ti plasmids are stably associated with their bacterial hosts. The possible role of the IS866 and IS867 elements in Ti plasmid evolution is discussed.
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  • 31
    ISSN: 1617-4623
    Keywords: Agrobacterium ; 6b gene ; Agrobacterium host range ; Oncogenes ; Plant growth regulation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The T-region located 6b gene of Agrobacterium tumefaciens has been found to interfere with cytokinin effects produced by the cotransferred ipt gene. We have compared the biological activity of three different 6b genes: A-6b from Ach5 (octopine, biotype 1), C-6b from C58 (nopaline, biotype 1) and T-6b from Tm4 (octopine, biotype III) by using different biological assays. Each 6b gene was inserted into a disarmed vector and tested on tobacco stems in coinfection experiments with the Ach5 cytokinin (ipt) gene (A-ipt). A-ipt/C-6b coinfections produced tumours with shoots, A-ipt/A-6b coinfections green tumours and A-ipt/T-6b coinfections tumours with a necrotic surface. The tumour phenotypes obtained were independent of the 6b/A-ipt coinfection ratios, indicating that the strain-specific 6b effects result from the activity of a non-diffusible 6b encoded product. Studies with ipt-less Tm4 mutants showed that 6b genes affect other tumour genes besides the ipt gene and pointed to an influence of T-6b on auxin effects resulting from the Tm4 iaa system. T-iaa/T-6b coinfection experiments showed that T-6b did indeed strongly increase tumour formation by the Tm4 iaa genes. The three 6b genes also have effects which do not require other T-region genes. The complex role of the 6b gene in crown gall induction and Agrobacterium host range will be discussed.
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  • 32
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    Molecular genetics and genomics 220 (1989), S. 65-68 
    ISSN: 1617-4623
    Keywords: Transformation ; Single-stranded DNA ; Agrobacterium ; Transient expression ; Extrachromosomal
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The transfer of the Agrobacterium T-DNA to plant cells involves the induction of the Ti plasmid virulence genes. This induction results in the generation of linear single-stranded (ss) copies of the T-DNA inside Agrobacterium and such molecules might be directly transferred to the plant cell. A central requirement of this ss transfer model is that the plant cell must generate a second strand and integrate the resulting double-stranded (ds) molecule into its genome. Here we report that incubating plant protoplasts with ss or ds DNA under conditions favouring DNA uptake results in transformation. The frequencies of transformation are similar and analysis of ss transformants suggests that the introduced DNA becomes double stranded and integrated. Analysis of transient expression from introduced ss DNA suggests that generation of the second strand is rapid and extrachromosomal.
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  • 33
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    Journal of Cellular Biochemistry 41 (1989), S. 179-188 
    ISSN: 0730-2312
    Keywords: protein kinase C ; TPA ; cell transformation ; gene expression ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: We have used a previously described retroviral expression vector pMV7-PKCβ1 to develop derivatives of two rat liver epithelial cell lines, K16 and K22, that stably express about tenfold-higher PKC activity than control cells. Despite these high levels of PKC, these cells did not exhibit gross morphologic changes, anchorage-independent growth, or tumorigenicity. K16PKC-4 and K22PKC-2, two lines with the highest PKC enzyme activity, were studied further in terms of several responses to the phorbol ester tumor promoter TPA. When treated with 100 ng/ml of TPA, the control K16MV7 and K22MV7 cells displayed a slight change in morphology, whereas the K16PKC-4 and K22PKC-2 cells displayed a marked change in morphology. Northern blot analyses demonstrated that TPA induced increased levels of fos, myc, phorbin, and ODC RNAs in control K16MV7 and K22MV7 cells, with maximum induction occurring at about 0.5, 1, 8, and 8 h, respectively. In K16PKC-4 and K22PKC-2 cells, TPA induction of phorbin and ODC RNAs was markedly enhanced, but this was not the case for myc and fos RNAs. In addition, the levels of myc RNA were constitutively higher in both K16PKC-4 and K22PKC-2 cells than in the control cells. Taken together, these results provide direct evidence that PKC plays a critical role in modulating the expression of myc, phorbin, and ODC RNAs. On the other hand, overexpression of PKCβ1 is not itself sufficient to cause cell transformation.
    Additional Material: 3 Ill.
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  • 34
    ISSN: 1432-2048
    Keywords: Agrobacterium ; Auxin in tumors ; Cytokinin (in tumors, turnover, oxidase) ; Mutant (T-DNA) ; Nicotiana (crown gall) ; T-DNA mutants
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The levels of the major cytokinins, zeatin, zeatin riboside, zeatin riboside-5′-monophosphate and zeatin-7-glucoside were measured in tobacco (Nicotiana tabacum L.) crown-gall tissues carrying insertion and deletion mutations in the T-DNA. Measurements were made by combined gas chromatography-mass spectrometry using selected ion monitoring with 15N- and 2H-labelled internal standards. The results demonstrate that, relative to wild-type tumour tissue, cytokinin levels are considerably elevated in tissues lacking functional T-DNA auxin-biosynthetic genes. From a detailed analysis of the major cytokinin metabolites it is concluded that a reduction in the extent of cytokinin degradation via N6-side-chain cleavage is an important factor leading to increased cytokinin levels in these tissues.
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  • 35
    ISSN: 1432-2048
    Keywords: Agrobacterium ; Co-inoculation ; Medicago ; Mutant (Rhizobium) ; Nodulation ; Rhizobium ; Root nodule initiation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Nodule formation on alfalfa (Medicago sativa L.) roots was determined at different inoculum dosages for wild-typeRhizobium meliloti strain RCR2011 and for various mutant derivatives with altered nodulation behavior. The number of nodules formed on the whole length of the primary roots was essentially constant regardless of initial inoculum dosage or subsequent bacterial multiplication, indicative of homeostatic regulation of total nodule number. In contrast, the number of nodules formed in just the initially susceptible region of these roots was sigmoidally dependent on the number of wild-type bacteria added, increasing rapidly at dosages above 5·103 bacteria/plant. This behavior indicates the possible existence of a threshold barrier to nodule initiation in the host which the bacteria must overcome. When low dosages of the parent (103 cells/plant) were co-inoculated with 106 cells/plant of mutants lacking functionalnodA, nodC, nodE, nodF ornodH genes, nodule initiation was increased 10- to 30-fold. Analysis of nodule occupancy indicated that these mutants were able to help the parent (wild-type) strain initiate nodules without themselves occupying the nodules. Co-inoculation withR. trifolii orAgrobacterium tumefaciens cured of its Ti plasmid also markedly stimulated nodule initiation by theR. meliloti parent strain. Introduction of a segment of the symbiotic megaplasmid fromR. meliloti intoA. tumefaciens abolished this stimulation.Bradyrhizobium japonicum and a chromosomal Tn5 nod- mutant ofR. meliloti did not significantly stimulate nodule initiation when co-inoculated with wild-typeR. meliloti. These results indicate that certainnod gene mutants and members of theRhizobiaceae may produce extracellular “signals” that supplement the ability of wild-typeR. meliloti cells to induce crucial responses in the host.
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  • 36
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    Theoretical and applied genetics 76 (1988), S. 767-774 
    ISSN: 1432-2242
    Keywords: Solanum tuberosum ; Transformation ; Agrobacterium ; Ethylene ; Phosphinotricin
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Leaves of the in vitro grown potato cultivars ‘Bintje’, ‘Berolina’, ‘Desiree’, and ‘Russet Burbank’ were wounded and co-cultivated with Agrobacterium strains having chimeric bar and nptII genes on a disarmed T-DNA. Each leaf from these cultivars formed numerous calli on kanamycin-containing medium, and almost all calli regenerated shoots. For ‘Russet Burbank’, it was necessary to include AgNO3 in the medium to obtain efficient shoot regeneration. The transformed plants have one to a few copies of the T-DNA, show NPT-II and PAT activities, and are resistant to high doses of the commercial preparation of phospinotricin (glufosinate). Almost no somaclonal variation was detected in trans-genic plants.
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  • 37
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    Cellular and molecular life sciences 44 (1988), S. 52-55 
    ISSN: 1420-9071
    Keywords: Human sperm ; human protamine ; protamine 1 ; protamine 2 ; protamine 3 ; infertility ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Protamines were extracted from the sperm of fertile and infertile human males and the relative proportion of protamines 1, 2, and 3 were determined by scanning microdensitometry following electrophoresis of total protamine in polyacrylamide gels. The proportion of the three protamines was found to be similar in sperm obtained from different normal males. The distribution of protamines in sperm obtained from a select group of infertile males producing an elevated level of large sperm heads, in contrast, was different from that of the fertile males.
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  • 38
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    Cellular and molecular life sciences 44 (1988), S. 910-919 
    ISSN: 1420-9071
    Keywords: Heart embryology ; developmental biology ; differentiation ; morphogenesis ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The subject of heart development has attracted the interest of many embryologists over the last two centuries. As a result, the main morphologic features of the developmental anatomy of the heart are already well established. Although there are still some controversial points, and there is probably much descriptive work yet to be done, emphasis is currently being placed on developmental mechanisms rather than simply on descriptive facts. The availability of new techniques and the overall advances in biological research are placing heart embryology in a new perspective. Today, we do not simply ask whether one or another embryonic structure arises further right or further left; instead, we are studying how cells, tissues, and their microenvironment interrelate at the several levels of biological organization (from the gene upwards) so as to give rise to a mature organ with a distinct shape and well-established functions. This paper attempts to review some of the basic aspects of the developmental anatomy of the heart. Descriptive embryology is used here as a tool. Emphasis is placed on developmental mechanisms, and on the present knowledge of how these mechanisms are related to the structural development of the heart.
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  • 39
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    Cellular and molecular life sciences 44 (1988), S. 930-936 
    ISSN: 1420-9071
    Keywords: Cardiac development ; embryogenesis ; gene expression ; complementary DNA ; molecular methodologies ; myocardial contractility ; myosin ; atrial natriuretic factor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Cardiac development involves a complex integration of subcellular processes into multicellular and, finally, whole organ effects. Until recently it has been difficult to investigate the genetic control of this organ level differentiation of the heart. The proliferation of molecular biology methodologies has provided mechanisms to directly investigate the control of these processes. This article focuses on molecular lines of research on two key areas in cardiac development: the regulation of expression of sarcomeric contractile and regulatory proteins, and atrial natriuretic factor. Molecular approaches are described which have allowed investigators to begin to determine the tissue and stage-specific expression of genes, to locate those genes in the genome, determine their sequences, and to directly investigate the mechanisms controlling their expression.
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  • 40
    ISSN: 1573-4927
    Keywords: cat hemoglobins ; polymorphism ; DNA ; restriction endonuclease ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract The molecular basis for the genetic control of variable proportions of the two cat hemoglobins Hb A(α2β 2 A ) and Hb B (α2β 2 B ) was investigated. Ratios of Hb A/Hb B vary between 50/50 and 90/10 among members of the mongrel cat population, with clusters around 50, 35, and 10% Hb B. Genomic DNA from cats of 50/50, 70/30, and 90/10 phenotypes were cut by restriction endonucleasesHindIII,EcoRI,BamHI,Bg1II,and Pst1 and hybridized to a fragment of the human β-globin gene. The results of the Southern blots suggested a pattern of homozygote, heterozygote, homozygote for the respective cat phenotypes, 50/50, 70/30, and 90/10. Therefore, the cat hemoglobin polymorphism seems to result from the possible combinations of an allelic gene pair.
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  • 41
    ISSN: 1573-7241
    Keywords: Atrial natriuretic peptide ; cardiac myocytes ; gene expression ; hybridization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Summary Primary cultures of neonatal cardiocytes expreses the gene for atrial natriuretic peptide (ANP). In general the levels of expression follow the rank order: atrial 〉 ventricular ≫ nonmyocardial cells. Following the initial dispersion cardlocytes require 48 to 72 hours before ANP seeretion and ANP mRNA accumulation approach a new steadystate level. In situ hybridization analysis indicates that ANP gene expression is concentrated in a subpopulation of cardiocytes in both the atrial and the ventricular cell cultures. These findings suggest that these primary cultures may be of value in defining the faciors governing the expression of the ANP gene in the cardiac cell.
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  • 42
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    Journal of bioenergetics and biomembranes 20 (1988), S. 19-39 
    ISSN: 1573-6881
    Keywords: E. coli unc operon ; H+-ATPase ; subunit stoichiometry ; gene expression ; codon usage ; translational initiation ; Shine-Dalgarno sequence ; recombinant DNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract Theunc (or atp) operon ofEscherichia coli comprises eight genes encoding the known subunits of the proton-translocating ATP synthase (H+-ATPase) plus a ninth gene (uncI) of unknown function. The subunit stoichiometry of the H+-ATPase (α 3β3γ1δ1ε1a1b2c10–15) requires that the respectiveunc genes be expressed at different rates. This review discusses the experimental methods applied to determining how differential synthesis is achieved, and evaluates the results obtained. It has been found that the primary level of control is translational initiation. The translational efficiencies of theunc genes are determined by primary and secondary mRNA structures within their respective translational initiation regions. The respective rates of translation are matched to the subunit requirements of H+-ATPase assembly. Finally, points of uncertainty remain and experimental strategies which will be important in future work are discussed.
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  • 43
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    Photosynthesis research 18 (1988), S. 7-31 
    ISSN: 1573-5079
    Keywords: Nicotiana ; chloroplast genome ; photosynthesis ; NADH dehydrogenase ; protein structure ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In order to understand more fully chloroplast genetic systems, we have determined the complete nucleotide sequence (155, 844 bp) of tobacco (Nicotiana tabacum var. Bright Yellow 4) chloroplast DNA. It contains two copies of an identical 25,339 bp inverted repeat, which are separated by 86, 684 bp and 18,482 bp single-copy regions. The genes for 4 different rRNAs, 30 different tRNAs, 44 different proteins and 9 other predicted protein-coding genes have been located. Fifteen different genes contain introns. Twenty-two genes for components of the photosynthetic apparatus have so far been identified. Most of the genes (except the gene for the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase) code for thylakoid membrane proteins. Twenty of them are located in the large single-copy region and one gene for a 9-kd polypeptide of photosystem I is located in the small single-copy region. The gene for the 32-kd protein of photosystem II as well as the gene for the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase have strong promoters and are transcribed monocistronically while the other genes are transcribed polycistronically. We have found that the predicted amino acid sequences of six DNA sequences resemble those of components of the respiratory-chain NADH dehydrogenase from human mitochondria. As these six sequences are highly transcribed in tobacco chloroplasts, they are probably genes for components of a chloroplast NADH dehydrogenase. These observations suggest the existence of a respiratory-chain in the chloroplast of higher plants.
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  • 44
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    Photosynthesis research 19 (1988), S. 39-61 
    ISSN: 1573-5079
    Keywords: Bacteriochlorophyll-protein complex ; gene expression ; light-harvesting complex ; reaction center ; Rhodospirillaceae ; transcriptional control ; translational control
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The puf operon of the purple nonsulfur photosynthetic bacterium, Rhodobacter sphaeroides, contains structural gene information for at least two functionally distinct bacteriochlorophyll-protein complexes (light harvesting and reaction center) which are present in a fixed ratio within the photosynthetic intracytoplasmic membrane. Two proximal genes (pufBA) specify subunits of a long wavelength absorbing (i.e., 875 nm) light harvesting complex which are present in the photosynthetic membrane in ≃15 fold excess relative to the reaction center subunits which are encoded by the pufLM genes. This review summarizes recent studies aimed at determining how expression of the R. sphaeroides puf operon region relates to the ratio of individual bacteriochlorophyll-protein complexes found within the photosynthetic membrane. These experiments indicate that puf operon expression may be regulated at the transcriptional, post-transcriptional, translation and post-translational levels. In addition, this review discusses the possible role(s) of newly identified loci upstream of pufB which may be involved in regulating either synthesis or assembly of individual bacteriochrlorophyll-protein complexes as well as the pufX gene, the most distal genetic element within the puf operon whose function is still unknown.
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  • 45
    ISSN: 1573-5028
    Keywords: Agrobacterium ; gene expression ; legumin (Pisum) ; Nicotiana ; seed storage protein ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A 3.4-kilobase genomic DNA fragment from Pisum sativum L. containing the LegA gene, which encodes a major legumin storage protein, was transferred to Nicotiana plumbaginifolia using an Agrobacterium tumefaciens strain containing the Bin 19 binary vector system. Northern hybridisation analysis of legA-transformed plants demonstrated that legumin-specific RNA was present in developing seeds but not in developing leaves. Legumin protein was immunologically detected in the mature seeds of legA-transformed plants, and was present as the correct-size protein composed of disulphide-bonded polypeptides. It is concluded that the transferred pea genomic fragment contains all the information necessary for seed-specific expression of the legA gene, and for correct processing of the primary transcript and the precursor legumin protein.
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  • 46
    ISSN: 1573-5028
    Keywords: potato ; patatin ; proteinase inhibitor ; cloning ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Tuberization in potato is a complex developmental process involving the expression of a specific set of genes leading to the synthesis of tuber proteins. We here report the cloning and analysis of mRNAs encoding tuber proteins. From a potato tuber cDNA library four different recombinants were isolated which hybridized predominantly with tuber mRNAs. Northern blot hybridization experiments showed that three of them, pPATB2, p303 and p340, can be regarded as tuber-specific while the fourth, p322, hybridizes to tuber and stem mRNA. Hybrid-selected in vitro translation and nucleotide sequence analysis indicate that pPATB2 and p303 represent patatin and the proteinase inhibitor II mRNA respectively. Recombinant p322 represents an mRNA encoding a polypeptide having homology with the soybean Bowman-Birk proteinase inhibitor while p340 represents an mRNA encoding a polypeptide showing homology with the winged bean Kunitz trypsin inhibitor. In total, these four polypeptides constitute approximately 50% of the soluble tuber protein. Using Southern blot analysis of potato DNA we estimate that these mRNAs are encoded by small multigene families.
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  • 47
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    Plant molecular biology 11 (1988), S. 641-649 
    ISSN: 1573-5028
    Keywords: codon usage ; gene expression ; histone H3 gene ; Medicago sativa ; somatic embryos
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A histone H3 gene was isolated from a dicotyledonous plant, alfalfa (Medicago sativa). The sequence analysis of this gene revealed no obvious GC preference in its codon usage. Apart from containing most of the typical consensus sequences found in both animal and plant histone genes, the alfalfa H3 gene exhibits distinct structural features such as (1) the unusual location of two GATCC motifs in its 5′ flanking sequence, (2) the existence of a CGCGGATC on the nonsense strand at position −232, (3) the existence of a long palindromic structure, and (4) several polyadenylation signal-like sequences in the 3′ flanking region. There are about 160 copies of histone H3 gene in alfalfa tetraploid genome. Using the alfalfa H3 gene as a probe to study the pattern of histone H3 transcripts in the alfalfa, we found that the H3 RNAs are undetectable in leaves, more in stems than in roots, and highest in somatic embryos. Moreover, the RNA products of H3 genes in all alfalfa tissues tested show unusually long nontranslated region compared to those of animal histone genes. An additional high molecular weight species of H3 transcript was detected only in somatic embryos.
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  • 48
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    Molecular and cellular biochemistry 84 (1988), S. 3-15 
    ISSN: 1573-4919
    Keywords: prostate ; androgens ; gene expression ; in situ hybridization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Summary After castration, the rat dorsolateral prostate M-40 mRNA initially decreased then rebounded to precastrated levels. The cellular site of M-40 expression and its renewed expression after castration was defined by in situ hybridization histochemistry. In situ hybridization with either a 32P-labeled or biotin-labeled M-40 cDNA probe demonstrated that M-40 mRNA levels were higher in the lateral than dorsal prostate. A second androgen regulated gene, RWB, also was highly expressed in the lateral prostate. The biotinylated cDNA probes provided microscopic resolution of the expressing cells, revealing two distinct morphologies of lateral epithelium which expressed both the M-40 and RWB mRNA. These morphologies appeared in ducts which contained either epithelial cell sheets that were highly convoluted or thinner epithelial cells with a minimal degree of convolution. The RWB mRNA decreased in both cell populations in response to androgen withdrawal. The decline and reappearance of M-40 mRNA also appeared in both epithelial cell types. These data demonstrated that after castration the M-40 mRNA initially decreased as expected for an androgen sensitive gene and then progressed to a fully inducible state. The mechanism of this progression remains to be elucidated.
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  • 49
    ISSN: 1573-5028
    Keywords: cDNA sequence ; chloroplast protein import ; gene expression ; glutamine synthetase ; nitrogen metabolism ; Phaseolus vulgaris L.
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Work using a full-length cDNA clone has revealed that the plastid-located glutamine synthetase (GS) of Phaseolus vulgaris is encoded by a single nuclear gene. Nucleotide sequencing has shown that this cDNA is more closely related to a cDNA encoding the plastidic GS of Pisum sativum than to cDNAs encoding three different cytosolic GS subunits of P. vulgaris. The plastid GS subunits are initially synthesized as higher M r (47000) precursors containing an N-terminal presequence of about 50 amino acids which is structurally similar to the presequences of other nuclear-encoded chloroplast proteins. The precursor has been synthesized in vitro and is imported by isolated pea chloroplasts and processed to two polypeptides of the same size as native P. vulgaris chloroplast GS subunits (M r 42000). Experiments with fusion proteins show that the N-terminal 68 amino acids of this precursor allow the cytosolic GS subunit β also to be imported and processed by isolated chloroplasts. Polyadenylated mRNA specifically related to the plastidic GS gene is most highly abundant in chloroplast-containing organs (leaves and stems) but is also detectable in roots and nodules.
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  • 50
    ISSN: 1573-5028
    Keywords: glyphosate ; herbicide ; 5-enolpyruvylshikimic acid 3-phosphate (EPSP) synthase ; enzyme overproduction ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Cell cultures of Corydalis sempervirens adapted to growth in the presence of 5 mM glyphosate [N-(phosphonomethyl)glycine] display a 30- to 40-fold increase in the cellular content of 5-enolpyruvylshikimic acid 3-phosphate (EPSP) synthase, the target enzyme of the herbicide. Translatable mRNA activity as well as transcript levels for EPSP synthase were increased 8-to 12-fold in the adapted (glyphosate-tolerant) as compared to the non-adapted (glyphosate-sensitive) cultures. Northern blot analysis revealed a single 1.8 kb transcript after hybridization with an oligonucleotide probe deduced from the N-terminal amino acid sequence of the enzyme. No significant differences in the relative abundance of EPSP synthase-specific DNA sequences could be detected, however, in Southern and dot blot analyses of restricted DNA isolated from the two cultures. We conclude that the overproduction of EPSP synthase in glyphosate-tolerant C. sempervirens cells is not based on the amplification of the corresponding gene.
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  • 51
    ISSN: 1573-5028
    Keywords: geminivirus ; Agrobacterium ; tomato golden mosaic virus ; agroinoculation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have adapted the “agroinfection” procedure of Grimsley and co-workers [4,5] to develop a simple, efficient, reproducible infectivity assay for the insect-transmitted, split-genome geminivirus, tomato golden mosaic virus (TGMV). Agrobacterium T-DNA vectors provide efficient delivery of both components of TGMV when used in mixed inoculation of wild-type host plants. A greater increase in infection efficiency can be obtained by Agrobacterium delivery of the TGMV A component to “permissive” transgenic plants. These “permissive” plants contain multiple tandem copies of the B component integrated into the host genome. An inoculum containing as few as 2000 Agrobacterium cells can produce 100% infection under these conditions. Further, our results show that there is a marked effect of the configuration of the TGMV A components within the T-DNA vector on time of symptom development. We have also found that transgenic plants carrying tandem copies of the A component do not complement the B component. Possible mechanisms to explain these results and the potential use of this system to further study the functions of the geminivirus components in infection are discussed.
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  • 52
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    Plant molecular biology 11 (1988), S. 495-506 
    ISSN: 1573-5028
    Keywords: abscisic acid ; barley aleurone layers ; cDNA cloning ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Abscisic acid (ABA) inhibits the gibberellic acid induced synthesis of α-amylase in barley aleurone layers, yet ABA itself induces more than a dozen polypeptides (Lin & Ho, Plant Physiol 82: 289–297, 1986). As part of our effort to elucidate the molecular action of ABA in barley aleurone layers, we have isolated and characterized an ABA-induced cDNA clone, pHV A1. This cDNA clone hybridizes to an RNA species of approximately 1.1 kb from ABA-treated barley aleurone layers. The level of this mRNA is tripled within 40 minutes after ABA treatment, reaches a peak at 8–12 h, and is present up to 48 h. The induction of this mRNA responds to concentrations of ABA as low as 10-9 M, but higher ABA concentrations induce higher expression of this mRNA. The products of hybrid-select translation and in vitro transcription/translation with pHV A1 comigrate on SDS gel as a 27 kDa polypeptide. However, the sequence of pHV A1 indicates that it has an open reading frame encoding a 22 kDa protein. This size discrepancy is probably due to the high content of the basic amino acid, lysine. This notion has been confirmed by two-dimensional gel electrophoresis showing that this polypeptide is one of the most basic proteins in ABA-treated barley aleurone layers. The deduced amino acid sequence of pHV A1 contains nine imperfect repeats 11 amino acids long which share homology with cotton Lea 7 protein (Baker, Steele & Dure, Plant Mol Biol, in press). The identity and function of the encoded product of pHV A1 is under investigation.
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  • 53
    ISSN: 1573-5079
    Keywords: Crassulacean acid metabolism ; gene expression ; Mesembryanthemum crystallinum ; mRNA levels ; soil salinity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Mesembryanthemum crystallinum responds to high salinity in the soil by shifting the mode of carbon assimilation from the C3 mode to Crassulacean acid metabolism (CAM). Several enzymes of carbon metabolism have increased apparent activities in the CAM mode, including phosphoenolpyruvate carboxylase (PEPcase) and pyruvate orthophosphate dikinase (PPDK). We have identified cDNA clones for PEPcase and PPDK by immunological screening of a cDNA library constructed in the protein expression vector lambda gt11. The clones were characterized by immunoblotting and RNA blotting techniques. RNA blotting showed that during CAM induction the steady-state level of mRNAs for both PEP case and PPDK increased.
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  • 54
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    Photosynthesis research 18 (1988), S. 205-222 
    ISSN: 1573-5079
    Keywords: ATP synthase ; chloroplast ; evolution ; gene expression ; operon
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Three CF1 and three CF0 subunits of the chloroplast H+-ATP synthase are encoded on the chloroplast genome. The chloroplast atp genes are organized as two operons in plants but not in the green alga, Chlamydomonas reinhardtii. The atpBE or β operon shows a relatively simple organisation and transcription pattern, while the atpIHFA or α operon is transcribed into a large variety of mRNAs. The atp genes are related to those of cyanobacteria and, more distantly, to those of non-photosynthetic bacteria such as E. coli, suggesting a common origin of most F1F0 ATP synthase subunits. Both the chloroplast and cyanobacterial ATP synthases have four F0 subunits, not three as in the E. coli complex. The proton pore of the CF0 is proposed to be formed by the interaction of subunits III and IV.
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  • 55
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    Photosynthesis research 17 (1988), S. 145-157 
    ISSN: 1573-5079
    Keywords: Carbon fixation ; enzyme assembly ; gene expression ; recombinant DNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The synthesis in Escherichia coli of both the large and small subunits of cereal ribulose bisphosphate carboxylase/oxygenase has been obtained using expression plasmids and bacteriophages. The level and order of synthesis of the large and small subunits were regulated using different promoters, resulting in different subunit pool sizes and ratios that could be controlled in attempts to optimize the conditions for assembly. Neither assembly nor enzyme activity were observed for the higher plant enzyme. In contrast, cyanobacterial large and small subunits can assemble to give an active holoenzyme in Escherichia coli. By the use of deletion plasmids, followed by infection with appropriate phages, it can be demonstrated that the small subunit is essential for catalysis. However, the small subunit is not required for the assembly of a large subunit octomer core in the case of the Synechococcus enzyme; self-assembly of the octomer will occur in an rbcS deletion strain. The cyanobacterial small subunits can be replaced by wheat small subunits to give an active enzyme in Escherichia coli. The hybrid cyanobacterial large/wheat small subunit enzyme has only about 10% of the level of activity of the wild-type enzyme, reflecting the incomplete saturation of the small subunit binding sites on the large subunit octomer, and possibly a mismatch in the subunit interactions of those small subunits that do bind, giving rise to a lower rate of turnover at the active sites.
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  • 56
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    Photosynthesis research 19 (1988), S. 63-71 
    ISSN: 1573-5079
    Keywords: CO2 fixation ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract From studies conducted in both our laboratory and by Gibson, Tabita and colleagues, as well as drawing on the recent studies with Alcaligenes eutrophus, we describe two genetic regions which have been identified on the chromosome of Rhodobacter sphaeroides which code for a number of enzymes involved in CO2 fixation. One region was found to contain the genes coding for fructose 1,6-bisphosphatase (fbpA), phosphoribulokinase (prkA), a 37 kDa polypeptide (cfxA), and form I ribulose 1,5-bisphosphate carboxylase/oxygenase (rbcL, S). These genes appear to be expressed in the same transcriptional direction and are tandomly arranged. A second, apparently unlinked region of the chromosome contains a duplicate (with respect to functionality of gene products) but not identical set of these same four genes. Although the gene order in both regions is apparently identical, there is approximately 4 kb of DNA separating the 3′-end of prkB and the beginning of cfxB. The specific genetic organizations and proposed roles of these two genetic regions are discussed.
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  • 57
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    Photosynthesis research 19 (1988), S. 129-152 
    ISSN: 1573-5079
    Keywords: chloroplast biogenesis ; gene expression ; ribosomes ; transcription ; translation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Isolated chloroplasts show substantial rates of protein synthesis when illuminated. This ‘in organello’ protein synthesis system has been advantageously utilised to elucidate the coding capacity of chloroplast and the regulation of chloroplast genes. The system is also being used recently to transcribe and translate homologous and heterologous templates. In this mini-review, we attempt to critically ecaluate the available literature and present the current and the prospective lines of research.
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  • 58
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    Plant and soil 108 (1988), S. 179-184 
    ISSN: 1573-5036
    Keywords: Agrobacterium ; genomic rearrangements ; molecular genetics ; Rhizobium ; strains instability ; symbiosis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract We have used molecular genetics techniques to analyze the structural and functional organization of genetic information ofRhizobium phaseoli, the symbiont of the common bean plantPhaseolus vulgaris. As in otherRhizobium species, the genome consists of the chromosome and plasmids of high molecular weight. Symbiotic determinants, nitrogen fixation genes as well as nodulation genes, are localized on a single replicon, the symbiotic (sym) plasmid. Thesym plasmid of differentR. phaseoli strains was transferred to anAgrobacterium tumefaciens strain cured of its native plasmids. In all cases, Agrobacterium transconjugants able to nodulate bean plants were obtained. Some of the transconjugants had the capacity to elicit an effective symbiosis. The genome ofR. phaseoli is complex, containing a large amount of reiterated DNA sequences. In mostR. pahseoli strains one of such reiterated DNA families corresponds to the nitrogenase structural genes (nif genes). A functional analysis of these genes suggested that the presence of reiteratednif genesis is related to the capacity of fixing atmospheric nitrogen in the symbiotic state. The presence of several repeated sequences in the genome might provide sites for recombination, resulting in genomic rearrangements. By analyzing direct descendants of a single cell in the laboratory, evidence of frequent genomic rearrangements inR. phaseoli was found. We propose that genomic rearrangements constitute the molecular basis of the frequent variability and loss of symbiotic properties in different Rhizobium strains.
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  • 59
    ISSN: 1573-5028
    Keywords: barley ; chymotrypsin inhibitor ; gene expression ; endosperm
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Full-length cDNA clones for barley chymotrypsin inhibitor 1 (CI-1) have been isolated from an endosperm-specific cDNA library. Hybridization and nucleotide sequence analyses indicate that these cDNAs represent two distinct types of CI-1 mRNA which we have called CI-1A and CI-1B. Both mRNAs encode polypeptides of 83 residues (M r=8790 and 8960) which differ at eleven positions. The full-length cDNA sequences do not predict N-terminal signal peptide extensions indicating that CI-1 is synthesized in the mature form in contrast to the homologous proteinase inhibitors of tomato and potato. Northern hybridization experiments show that the CI-1 genes are under strict developmental and organ-specific control. CI-1 transcripts were first detected in the developing barley endosperm between 12 and 14 days after anthesis but no CI-1-related sequences were detected in the RNA preparations from shoots, leaves or roots. The expression of CI-1 was also studied in the high-lysine barley mutants Hiproly, Risø 56 and Risø 1508. Approximately 15-fold (Hiproly) and 4-fold (Risø 56 and 1508) higher levels of CI-1 mRNA were detected in the mutant endosperms compared to normal barley. These results correlate well with the increased deposition of CI-1 in the high-lysine lines and show that the differential expression is controlled mainly at the level of transcription or stability of the mRNA. Using Southern-blots of barley DNA we estimate that there are three copies of CI-1 per haploid genome in both normal and mutant barley lines.
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  • 60
    ISSN: 1617-4623
    Keywords: Agrobacterium ; Crown gall ; Biological control ; Agrocin 84 ; Agrocin plasmid transfer
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Agrobacterium radiobacter strain K84 is used commercially for the biological control of crown gall. It contains the conjugative plasmid pAgK84, which encodes the synthesis of agrocin 84, an antibiotic that inhibits many pathogenic agrobacteria. A breakdown of control is threatened by the transfer of pAgK84 to pathogens, which then become resistant to agrocin 84. A mutant of pAgK84 with a 5.9-kb deletion overlapping the transfer (Tra) region was constructed using recombinant DNA techniques. The BamHI fragment B1 which covers most of the Tra region was cloned in pBR325 and its internal EcoRI fragments D1 and H, which overlap the Tra region, were removed, leaving 3.7 kb and 0.5 kb of pAgK84 on either side of the deletion. The latter was increased to 3.3 kb by adding EcoRI fragment D2 from a BamHI fragment C clone. The modified pBR325 clone was mobilized into Agrobacterium strain NT1 harbouring pAgK84 with a Tn5 insertion just outside the Tra region but covered by the deletion. A Tra+ cointegrate was formed between the Tn5-insertion derivative and the pBR325-based deletion construct by homologous recombination. The cointegrate was transferred by conjugation to a derivative of strain K84 lacking pAgK84, in which a second recombination event generated a stable deletion-mutant by deletion-marker exchange. The resultant new strain of A. radiobacter, designated K1026, shows normal agrocin 84 production. Mating experiments show that the mutant plasmid, designated pAgK1026, is incapable of conjugal transfer at a detectable frequency.
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  • 61
    ISSN: 1617-4623
    Keywords: Agrobacterium ; Small subunit of ribulose bisphosphate carboxylase ; Chlorophyll a/b binding protein ; Promoter comparisons ; Chitinase activity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A bacterial chitinase gene from Serratia marcescens (chiA) was fused to (i) a promoter of the ribulose bisphosphate carboxylase small subunit (rbcS) gene and (ii) two different chlorophyll a/b binding protein (cab) gene promoters from petunia. The resulting constructions were introduced into Agrobacterium Ti plasmid-based plant cell transformation vectors and used to generate multiple independent transgenic tobacco plants. ChiA mRNA and protein levels were measured in these plants. On average, the rbcS/chiA fusion gave rise to threefold more chiA mRNA than either cab/chiA fusion. We investigated the influence of sequences around the translational initiation ATG codon on the level of ChiA protein. The rbcS/chiA and cab/chiA fusions in which the sequence in the vicinity of the translational initiation codon is ACC ATGGC gave rise to transformants with higher levels of ChiA protein than those carrying a cab/chiA fusion with the sequence CAT ATGCG in the same region. This difference in translational efficiency is consistent with previous findings on preferred sequences in this region of the mRNA. In those transformants showing the highest level of ChiA expression, ChiA protein accumulated to about 0.25% of total soluble leaf protein. These plants contained significantly higher chitinase enzymatic activity than control plants.
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  • 62
    ISSN: 1617-4623
    Keywords: Promoter strength ; Transformed callus ; Chitinase gene ; Octopine synthase gene ; Agrobacterium
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The 35S promoter of cauliflower mosaic virus and promoters from the nopaline synthase, 1′ and 2′ genes of Agrobacterium tumefaciens T-DNA were fused to the bacterial octopine synthase and chitinase gene coding regions. These chimaeric gene constructions were introduced into tobacco, sugarbeet and oilseed rape cells and their relative levels of expression measured by primer extension analysis of RNA isolated from pooled populations of stably transformed calli. In tobacco callus, the 35S promoter provided the highest levels of gene expression, followed by the 2′, 1′ and nopaline synthase promoters. While the ranking of these promoters is conserved in sugarbeet and oilseed rape callus, there is between-species variation in the relative strength of these promoters. In all three species, transcription initiation is conserved for each of the chimaeric gene constructions. Additional constructions in which the 5′ untranslated leader of a petunia chlorophyll a/b binding protein gene is substituted for DNA downstream of the 35S transcription start site demonstrates that heterologous 5′ leader sequences can be utilized to augment steady-state levels of reporter gene expression
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  • 63
    ISSN: 1573-0778
    Keywords: CG suppression ; DNA methylation ; gene expression ; MHC ; mutation hot spots
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract DNA methylation of human class II genes of the Major Histocompatibility Complex (MHC) was correlated with gene expression and methyl-related CG → TG and CG → CA changes. It was found that cytosine methylation of the CG dinucleotides of MHC class II genes should be involved in generating a fraction of nucleotide polymorphism, rather than in controlling transcription.
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  • 64
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    New York, N.Y. : Wiley-Blackwell
    Journal of Cellular Biochemistry 38 (1988), S. 205-212 
    ISSN: 0730-2312
    Keywords: cytokines ; gene expression ; retroviral promoters ; antitumoral activities ; phenotypical reversion ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: The potential mechanisms of interferon (IFN)-gamma and tumor necrosis factor (TNF)-alpha action on tumor cells have been investigated in a model of mouse fibroblasts transformed by distinct retroviral vectors carrying the v-mos, c-myc, and v-Ha-ras oncogene, respectively. Treatment with both cytokines not only caused growth inhibition of v-mos- and c-myc-transformants, but also a reversion of transformation-induced suppression of major histocompatibility complex (MHC) class I antigen expression in all transformed cell lines. The phenotypical reversion of transformants was preceded by a selective modulation of LTR-controlled oncogene expression. TNF-alpha primarily affected stability of oncogene-specific RNAs without influencing the activity of retroviral promoters. In contrast, IFN-gamma was effective at the transcriptional level, apparently due to inhibition of LTR activity as revealed from reduced CAT activity in IFN-gamma-treated LTR-CAT transformants. This IFN-gamma-mediated down-regulation of retroviral promoter activity seemed to be selective for Moloney-virus-derived promoters, since the activity of other viral and cellular promoters was not suppressed by IFN-gamma.
    Additional Material: 1 Ill.
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  • 65
    ISSN: 0730-2312
    Keywords: ADP-ribosyl transferase ; cell proliferation ; gene expression ; protein kinase C ; c-myc expression ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Human α or β interferons inhibit the proliferation of Daudi Burkitt lymphoma cells and induce the differentiation of these cells towards a mature plasma cell phenotype. Similar responses are seen when Daudi cells are treated with the phorbol ester, TPA. Both interferons and TPA down-regulate expression of the c-myc oncogene in these cells. Although TPA can mimic the effect of interferon on cell differentiation, it does not induce 2′5′ oligoadenylate synthetase or the interferon-sensitive mRNAs, 6-16 or 9-27. Thus chronic stimulation of protein kinase C by TPA cannot mimic all of the effects of interferon treatment on gene expression. Inhibition of ADP-ribosyl transferase activity by 3-methoxybenzamide impairs interferon- or TPA-induced differentiation of Daudi cells. This agent induces a higher level of c-myc mRNA in the cells and stimulates the incorporation of [3H]thymidine into DNA; although these effects are partially counteracted by interferon or TPA treatment, the elevated expression of the c-myc gene may be sufficient to prevent terminal differentiation and allow cell proliferation to continue.
    Additional Material: 3 Ill.
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  • 66
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    Journal of Cellular Biochemistry 37 (1988), S. 243-253 
    ISSN: 0730-2312
    Keywords: hepatocyte differentiation ; gene expression ; hormonal control ; glucocorticoids ; insulin ; cyclic AMP ; tyrosine aminotransferase ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Cloned cDNAs were used in hybridization analyses to assess hormonal responsiveness of two similarly regulated genes in livers of late-term fetal rats. Transcription of the tyrosine aminotransferase gene and of gene 33 (Lee et al.: J Biol Chem 260:16433-16438, 1985) is enhanced by glucocorticoids and by each of the usually antagonistic hormonal agents, insulin and cAMP, in adult liver, and that of both genes is developmentally activated at or just prior to birth. The mRNA of gene 33 was found to be significantly increased by each of the hormonal regulators in livers of fetuses treated in utero. Expression of the nearly silent aminotransferase gene in fetal liver was appreciably increased by cAMP but was refractory to control by either glucocorticoids or insulin; capacity of this gene to respond to insulin was not realized until several days postpartum. The data indicate specificity in the developmental acquisition of the capacity of individual genes to respond to hormonal regulators.
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  • 67
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 9 (1988), S. 167-180 
    ISSN: 0192-253X
    Keywords: enzyme pattern ; gene expression ; protein synthesis ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Aldehyde oxidase (AO) and pyridoxal oxidase (PO) distribution patterns were determined in the imaginal wing discs for a series of strains of Drosophila melanogaster heterozygous for different Minute mutations. The mutant severity ranged from very weak to strong. The results show an inverse response of AO and PO to the expressivity of the Minute mutation: in weaker Minutes the extent of the AO positive area increases, whereas PO activity disappears. The results are discussed with reference to an impaired protein synthesis in Minutes.
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  • 68
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    Developmental Genetics 9 (1988), S. 13-22 
    ISSN: 0192-253X
    Keywords: ontogeny ; lethality ; gene expression ; mRNA ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Cadmium is a toxic metal that induces the expression of metallothionein genes in many tissues and that binds avidly to metallothionein, a soluble transition metal binding protein. The present study examined the temporal pattern and magnitude of accumulation of metallothionein mRNA in liver of C57BL/6J mice of various ages treated with cadmium. In adult female mice, accumulation was dependent on the dosage level of cadmium and related to the concentration of this metal in liver. The accumulation of metallothionein mRNA in liver depended on age at exposure to cadmium. Intraperitoneal administration of 2 mg of cadmium per kg provoked small increases (two- to threefold) in levels of metallothionein mRNA in livers of 7- and 14-day-old mice. In contrast, cadmium treatment of 28- and 56-day-old mice resulted in 12- to 19-fold increases in levels of metallothionein mRNA in liver with maximum increases occurring 3 to 4 hr after treatment. Because similar patterns for the accumulation of cadmium of liver were found in 7-, 28-, and 56-day-old mice, observed age-dependent differences in induction of metallothionein mRNA in liver were probably not due to differences in the accumulation of cadmium in this organ. Taken together, these data suggest that tissue-specific factors controlling the expression of metallothionein genes may account for developmental variation in the inducibility of these genes by cadmium. Ontogenic variation in accumulation of metallothionein mRNA after cadmium treatment may be a factor in developmental variation in the acute lethality of cadmium in C57BL/6J mice.
    Additional Material: 5 Ill.
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  • 69
    ISSN: 0192-253X
    Keywords: Daucus carota ; auxin ; gene expression ; mutant ; filtration-enrichment procedure ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Cell cultures of the carrot Daucus carota are a useful experimental system for studying the genetic regulation of plant embryogenesis. A modified filtration-enrichment procedure was used to isolate 21 temperature-sensitive variants in somatic embryogenesis; the variants display normal embryo development at the permissive temperature (24°C) and altered development at the restrictive temperature (33°C). Temperature-shift experiments were performed on these variants to determine the timing of gene action for the putative temperature-sensitive alleles. According to their phenotypes at the restrictive temperature, these variants can be divided into six classes: No Growth, Callus Proliferation, Globularstage Block, Oblong-stage Block, Lateral Growth, and Root Formation. Although many variants exhibit lengthy temperature-sensitive periods, the temperature sensitivity of some variants is restricted to one or two embryonic stages. These results plus those in the literature are incorporated into a preliminary model concerning the genetic regulation of carrot embryogenesis.
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  • 70
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 9 (1988), S. 259-265 
    ISSN: 0192-253X
    Keywords: G-proteins ; gene expression ; developmental regulation ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: We have identified a two-member gene family in the Dictyostelium genome and have isolated corresponding cDNA or genomic DNA recombinant clones. Analyses of these DNA sequences predicted encoded proteins of ∼200 amino acids with ∼90% sequence identity to each other. These Dictyostelium proteins also share amino acids identity within the GTP-binding domains in the family of G-regulatory proteins involved in cellular regulation and transmembrane signalling. Additional structural similarities are seen with members of the ras supergene family, such as ras, ral, and rho. They are similar in size (usually ∼200 amino acids), possess four conserved domains involved in GTP interaction and are believed to be anchored in the membrane by fatty acid modification of a cysteine residue near the carboxy terminus. More extensive identity is observed with YPT1 and SEC4, two other members of this family of genes that are essential in yeast. The amino-terminal half of both Dictyostelium proteins is 70% identical in amino acid sequence to the YPT1 and SEC4 yeast proteins with less identity continuing through the remainder of the proteins. In addition these proteins terminate in two cysteine residues that are thought to be required for membrane anchorage.The two genes within this Dictyostelium family are organized differently in the genome and are differentially regulated during development. One gene is colinear in sequence with its mRNA in the protein coding region, whereas the other gene encodes a spliced mRNA. The intron-containing gene is associated with a developmentally regulated (AAC)-repeat sequence. Finally, we have shown that the expression of one of the genes is induced during development with kinetics similar to that of other (AAC)n-associated genes; conversely, the expression of the second gene is repressed at a similar developmental stage.
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  • 71
    ISSN: 0192-253X
    Keywords: cAMP ; Ca2+ ; signal transduction ; cell surface receptor ; gene expression ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Elevated levels of cAMP are essential for the expression of many postaggregation prespore and prestalk mRNA species and for the suppression of some growth phase mRNAs. Here we review evidence that this regulation is mediated by cAMP interacting at the cell surface receptor. These effects of cAMP on gene expression can occur under conditions where the receptor-associated adenylate cyclase is inactivated and in concentrations that are consistent with receptor binding. A number of differences are noted in the mechanism by which cAMP regulates prespore and prestalk genes. Finally, evidence is reviewed for the role of a Ca2+-dependent signal transduction system in coupling the expression of some of the prespore mRNAs to the cAMP receptor. This signal transduction system does not appear to be involved in the expression of the cAMP-dependent prestalk gene.
    Additional Material: 2 Ill.
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  • 72
    ISSN: 1432-2048
    Keywords: Agrobacterium ; Cytokinin biosynthesis ; Auxin autonomy ; Nicotiana (auxin autonomy) ; Tumor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Agrobacteria carrying mutations at the auxin-biosynthesizing loci (iaaH and iaaM of the Ti plasmid) induce shoot-forming tumors on many plant species. In some cases, e.g. Nicotiana glutinosa L., tumors induced by such mutant strains exhibit an unorganized and fully autonomous phenotype. These characteristics are stable in culture at both the tissue and cellular level. We demonstrate that the cytokinin-biosynthesis gene (ipt) of the Ti plasmid is responsble for the induction of both auxin and cytokinin autonomy in N. glutinosa. Cloned cell lines carrying an ipt gene but lacking iaaH and iaaM are capable of accumulating indole-3-acetic acid. Interestingly, non-transformed N. glutinosa tissues exhibit an auxin-requiring phenotype when they are grown on medium supplemented with an exogenous supply of cytokinin. These results strongly indicate that exogenously supplied cytokinin does not mimic all the effects of the expression of the ipt gene in causing the auxin-autonomous growth of N. glutinosa cells.
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  • 73
    ISSN: 1432-2048
    Keywords: Agrobacterium ; Crown-gall ; DNA, transferred ; Nicotiana (T-DNA) ; T-DNA structure
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Data are provided which show that transferred DNA (T-DNA) present in Nicotiana plumbaginifolia crown-gall lines in most cases was scrambled and not intact. Both wild-type, and ‘rooter’- and ‘shooter’-type mutants of octopine-type Agrobacterium tumefaciens were used to infect N. plumbaginifolia plantlets, cultured in vitro. Resulting tumors were excised from the plantlets and cultured for more than three years. During subculturing the tumor lines were scored for the following phenotypic traits: phytohormone autonomous growth in vitro (Aut+), spontaneous shoot regeneration (Reg+), root deficiency of shoots (Rod+), octopine production (Ocs+) and mannopine and agropine production (Mas+Ags+). An unexpectedly large variety of phenotypes was observed. For instance, two out of three tumor lines induced on haploid plantlets by the rooter mutant LBA4210 regenerated shoots, a phenomenon which is not observed for octopine tobacco tumor lines. Fifty percent of the crown-gall lines studied did not contain octopine. Only one line out of six independent lines analyzed was found to have a ‘regular’ T-DNA structure. Occurrence of aberrant T-DNA structures was not correlated with the ploidy level of infected plantlets, nor with the T-region structure of the inciting bacterial strain. The pattern of TL-DNA transcripts was studied for one line and correlated well with the aberrant T-DNA structure detected. Segments of TR-DNA, having irregular structures as well, were detected in two out of the six lines studied. The scrambled nature of the TR-DNA explained the absence of mannopine and agropine in these two lines. In addition, it was observed that N. plumbaginifolia tissue lines which did not carry T-DNA, became readily phytohormone autotrophic (habituated) at an early stage in tissue culture.
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  • 74
    ISSN: 1432-2048
    Keywords: Agrobacterium ; Auxin biosynthesis ; Indole-3-acetamide ; Pseudomonas ; Tryptophan-2-mono-oxygenase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract It has been proposed that the “eukaryotic” T-DNA-encoded indole-3-acetic acid (IAA) biosynthesis genes of Agrobacterium tumefaciens and their prokaryotic counterpart in Pseudomonas savastanoi originated from common ancestor genes. This paper provides additional evidence for the functional similarity between the gene products. We have demonstrated that a chimeric gene consisting of the coding sequence of the P. savastanoi tryptophan-2-mono-oxygenase (iaaM gene) and a plant promoter encodes an active enzyme in Nicotiana tabacum. Transformants obtained with this chimeric gene grew as a callus on hormone-free media. No stably transformed plantlets could be isolated. The callus tissues contained extremely high levels of indole-3-acetamide and slightly elevated levels of IAA. Either indole-3-acetamide by itself has a low auxin activity or, alternatively, it is converted aspecifically and at low rates into IAA. The P. savastanoi tryptophan-2-mono-oxygenase activity in plants is also able to detoxify the amino-acid analogue 5-methyltryptophan. This property can be used for positive selection of transformed calli.
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  • 75
    ISSN: 1432-2048
    Keywords: Agrobacterium ; Crown gall ; Nicotiana (crown gall, T-DNA) ; Transformation (tobacco) ; Transferred DNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Grafts from the SR1 tobacco crown-gall lines NT1 (having a deletion eliminating part of the transferred (TL)-DNA auxin locus) and NT2 (having an IS60 insertion in gene 2 of this auxin locus) were cross-pollinated with pollen from nontransformed SR1 tobacco plants. One half of the resulting F1 progeny resembled the female parent (“transformed” NT1-like and NT2-like seedlings respectively) and one half resembled the male parent (“non-transformed” SR1-like seedlings). For three states of differentiation (callus, shoot, graft) all phenotypic markers of the transformed seedlings studied were identical to those of the transformed female parent. Most phenotypic markers of non-transformed seedlings corresponded with markers of the male parent. Unlike the SR1 male parent, however, the SR1-like seedlings showed the maternal traits hyperstyly and male sterility. These two traits were inherited by 100% of the F1 seedlings studied. Ninety percent of the non-transformed F2 seedlings were still male-sterile whereas in as much as 50–100% of the non-transformed F3 progeny, male fertility had been restored. The SR1-like F1 seedlings did not contain any T-DNA. At the level of restriction-fragment analysis the T-DNA structures of all 22 NT1-like seedlings examined were identical to the T-DNA structure of their female parent NT1. The steady-state level of transcripts 4 (cytokinin locus) and 6a/6b relative to transcript 3 (octopine-synthase locus) was less in shoots and grafts than in callus. Observed variation in shoot morphology among the twenty-two NT1-like seedlings was not correlated with T-DNA structure, organization and expression at the level of steady-state mRNA. The T-DNA structure of NT2 and its transformed seedlings deviated from regular border-to-border TL-DNA, in that it extended beyond the left border repeat.
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  • 76
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    Cellular and molecular life sciences 43 (1987), S. 741-750 
    ISSN: 1420-9071
    Keywords: Hybridization histochemistry ; hybridocytochemistry ; in situ hybridization ; gene expression ; histochemical hybridization ; nucleic acid hybridization ; histocytochemistry
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The location of gene expression by hybridization histochemistry is being applied in many areas of research and diagnosis. The aim of this technique is to detect specific mRNA in cells and tissues by hybridization with a complementary DNA or RNA probe. Requirements for optimal specificity, sensitivity, resolution and speed of detection may not all be encompassed in one simple technique suitable for all applications, thus appropriate procedures should be selected for specific objectives. With reference to published procedures and our own extensive experience, we have evaluated fixatives, probes, labels and other aspects of the technique critical to the preservation and hybridization in situ of mRNA and detection and quantition of hybrids.
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  • 77
    ISSN: 1572-994X
    Keywords: EBV ; gene expression ; immortalization ; cDNA cloning ; Northern blot hybridization ; tonsil lymphocytes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Two infection systems have been introduced in order to study viral gene expression at the initial period of Epstein-Barr virus (EBV) infection which leads to immortalization. The data indicate that major viral gene expression in tonsil lymphocytes at 2 days post-infection (p.i.) with EBV is very similar to that observed in latently infected cells. Both tonsil lymphocyte and BJAB cell (lymphoblastoid cells free of EBV genome) infection with EBV induced similar viral gene transcription. Twelve cDNA clones were prepared from poly(A) RNA of tonsil lymphocytes infected with EBV 2 days p.i. by hybridization with BamHI fragments of EBV DNA. Some cDNAs were derived from primary transcripts of the BamHI-WYHK region, suggestive of splicing of a large transcript. It is possible that a number of cDNA clones may be derived from cellular genes. The derivation of these cDNA clones is being studied.
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  • 78
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    Plant molecular biology 8 (1987), S. 251-264 
    ISSN: 1573-5028
    Keywords: chromatin structure ; DNase I hypersensitivity ; gene expression ; sucrose synthetase ; Zea mays
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The local chromatin structure of the Shrunken-1 (Sh) gene of maize was probed by analyzing DNase I hypersensitivity. Sh encodes the gene for sucrose synthetase, a major starch biosynthetic enzyme, which is maximally expressed in the endosperm during seed maturation. In addition to general DNase I sensitivity, specific DNase I hypersensitive sites were identified in endosperm chromatin that mapped near the 5′ end of the Sh gene. The pattern of hypersensitive sites and their relative sensitivity were altered in other non-dormant tissues that produce little or no enzyme. However, some changes in chromatin structure appear to be independent of Sh gene expression and may reflect general alterations associated with plant development. The chromatin structure of several sh mutations, induced by Ds controlling element insertions, was also analyzed. Although the insertions perturbed expression of the gene, there were no notable effects on local chromatin structure.
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  • 79
    ISSN: 1573-5028
    Keywords: potato tuber ; elicitor ; arachidonic acid ; cDNA cloning ; gene expression ; PR proteins
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Accumulation of messenger RNAs in potato tuber discs was analysed during the hypersensitive response induced by treatment with the biotic elicitor arachidonic acid. In vitro translation of polysomal poly(A)+ RNAs indicated that the accumulation of some sixteen mRNAs varied following treatment with arachidonic acid, and that the level of thirteen of these was increased. Two cDNA closes (pSTH-1 and-2) were isolated from a library of elicitor-treated tissue cDNAs. Northern blot analysis using these clones as molecular probes indicated that the levels of at least two mRNAs were markedly increased after elicitor treatment. In hybrid-released translation experiments, each of the cDNA clones selected more than one mRNA. Translation of these mRNAs yielded two polypeptides of Mr 45 000 (for the pSTH-1 clone), and three polypeptides of Me 17 000 (for the pSTH-2 clone). The low molecular weight polypeptides may correspond to potato pathogenesis-related (PR) proteins.
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  • 80
    ISSN: 1573-5028
    Keywords: Agrobacterium ; nodulin gene expression ; Rhizobium ; root nodule ; sym-plasmid ; Vicia
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Nodulin gene expresison was studied in Vicia sativa (common vetch) root nodules induced by several Rhizobium and Agrobacterium strains. An Agrobacterium transconjugant containing a R. leguminosarum symplasmid instead of its Ti-plasmid, that was previously shown to form “empty” nodules on pea, induced nodules on Vicia roots in which nodule cells were infected with bacteria. In the Vicia nodules induced by this transconjugant, two so-called early nodulin genes were found to be expressed, whereas in the nodules formed on pea the expression of only one early nodulin gene was detected. In both cases the majority of the nodulin genes was not expressed. Apparently, an intracellular location of the bacteria is not sufficient for the induction of the majority of the nodulin genes. All nodulin genes were expressed in nodules induced by cured Rhizobium strains containing cosmid clones that have a 10 kb nod region of the sym-plasmid in common. Since in tumours no nodulin gene expression was found at all, the Agrobacterium chromosome does not contribute to the induction of nodulin genes. Therefore it is concluded that the signal for the induction of the expression of the two Vicia early nodulin genes is encoded by the nod-region, and the signal involved in the induction of all other nodulin genes has to be located outside the sym-plasmid, on the Rhizobium chromosome. The apparent difference in early nodulin gene expression between pea and Vicia is discussed in the light of the usefulness of Agrobacterium transconjugants in the study of nodulin gene expression.
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  • 81
    ISSN: 1573-5028
    Keywords: azacytidine ; DNA/RNA hybridization ; gene expression ; marker rescue ; nopaline-synthase ; plant tumor cells ; Ti-plasmid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Cell wall regenerating protoplasts from soybean cells kept in suspension culture were cocultivated with bacteria which were derived from the nopaline strain C58 of Agrobacterium tumefaciens. When the bacteria carried an oncogenic Ti-plasmid, about 5% of the surviving protoplasts were able to form calli on hormone-free agar in contrast to controls, where bacteria without Ti-plasmid were applied, and where no calli were formed. After isolation of DNA from hormone-independently growing cells further evidence for transformation was obtained by hybridization to Ti-plasmid specific RNA and by rescue of a segment with a bacterial resistance gene which had been inserted before into the T-DNA. Transfer of T-DNA harboring a neomycin-resistance gene activated by the nos-promoter resulted in calli growing on kanamycin. Verification of segments located at the left and the right part of the T-DNA indicated the presence of its entire length in transformed soybean cells. Expression of T-DNA genes was measured by the assay of nopaline-synthase. Cells cultured on agar had a much higher level of nopaline-synthase than fast growing cells in suspension culture. Transferring them to agar or treatment with azacytidine strongly increased synthesis of nopaline-synthase indicating a reversible repression presumably via a methylation mechanism.
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  • 82
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    Plant molecular biology 9 (1987), S. 345-375 
    ISSN: 1573-5028
    Keywords: Patatin ; Solanum tuberosum L. ; Agrobacterium ; chimaeric gene ; chloramphenicol acetyltransferase (CAT)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A member of the patatin multigene family which encodes the major soluble tuber protein was isolated from potato cultivar Désirée. Analysis by strategic nucleotide sequencing demonstrated close homology to analogous regions of previously isolated patatin genomic clones from different cultivars. A 3.8-kb fragment containing the promoter and 5′ flanking DNA of the patatin gene was used to construct a transcriptional fusion gene with the coding DNA of the bacterial chloramphenicol acetyltransferase (CAT) gene and the polyadenylation/termination sequences of the nopaline synthase gene (nos). The chimaeric gene was reintroduced into potato cultivar Désirée by agrobacterial transformation of tissue slices. Regenerated transformed plants showed expression of the chimaeric gene (as determined by CAT activity) in tubers, but not in leaves, stems or roots of in vitro grown plants. Independent transformants did not show substantial variation in the level of induced tuber-specific CAT activity. Thus, information contained within 3.8 kb of the 5′ flanking DNA of the patatin gene analysed is sufficient to direct tuber-specific expression, largely independent of position effects.
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  • 83
    ISSN: 1573-5028
    Keywords: gene expression ; nitrogen fixation ; nodulins ; Phaseolus vulgaris ; Rhizobium ; root-nodule development
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The identification of some nodule-specific host proteins (nodulins) from common bean (Phaseolus vulgaris L.), a tropical ureide-transporting legume, is described. Particularly, the existence and developmental expression of several abundant nodule-specific transcripts of P. vulgaris are shown, including leghemoglobin, nodulespecific uricase and a group that in vitro translates into a cluster of about 30 kDa products. The expression pattern of nodulins in effective (Fix+) nodules compared to ineffective (Fix-) ones is also presented. The modified expression of main nodulins observed between these nodules indicates that different levels and/or factors associated with their regulation are involved. The intracellular infection by Rhizobium as a decisive step in the induction of some P. vulgaris nodulins is discussed.
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  • 84
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    Plant molecular biology 9 (1987), S. 547-563 
    ISSN: 1573-5028
    Keywords: acetate ; blue light ; Chlamydomonas reinhardtii ; chlorophyll a/b-binding protein gene ; gene expression ; LHCP gene
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In Chlamydomonas reinhardtii, the chlorophyll a/b-binding proteins of photosystem II are encoded in the nucleus by a small family of genes. We have studied the expression of one gene, which we call cabII-1, in a green-in-the-dark strain, which can synthesize chlorophyll in the dark or light, and in a yellow-in-the-dark mutant strain, which is able to make chlorophyll only in the light. In light/dark synchronized cultures of both strains, cabII-1 mRNA abundance increases during the first 6 h of a 12-h light phase, remains high for several hours, then declines. A variety of illumination conditions have been used to analyze the cabII-1 mRNA increase: continuous or intermittent red, blue, or white light, with or without 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), an inhibitor of photosystem II. Our results suggest that light induces increased cabII-1 transcript abundance in two ways: 1) by virtue of its role in the light reactions of photosynthesis and 2) by a blue lightstimulated mechanism which is independent of photosynthesis. We have also examined the role of acetate in regulating cabII-1 mRNA levels in the dark. In both green- and yellow-in-the-dark strains, 15 mM Na-acetate, added to synchronized cells in the dark, induces an increase in cabII-1 mRNA abundance with a temporal accumulation pattern very similar to that induced by continuous white light. We suggest that by providing an energy source, acetate stimulates cellular growth, cell cycle progression, and increased cabII-1 mRNA abundance. Interestingly, in cells exposed to light, acetate inhibits the light-induced increase in cabII-1 mRNA abundance by a mechanism which is not yet understood.
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  • 85
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    Bioscience reports 7 (1987), S. 955-963 
    ISSN: 1573-4935
    Keywords: gene expression ; GC box ; negative regulation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract A DNA control sequence T G GGGCGG AAT GGC , or the “GC” box, has been described in the promoter regions upstream of a number of eukaryotic genes transcribed by polymerase II (for review, see Dynan, W. S. and Tjian, R.,Nature 316:774, 1985). The “GC” box can occur in single or multiple copies and is the binding site for a protein factor, Spl, which activates initiation of transcription. We have observed in the rainbow trout protamine gene 3′ to the TATA box, three “GC” boxes spaced at 80 bp intervals. The first is 5′ to the cap site and possesses the ability to “silence” transcription from the protamine promoter in constructs linking this promoter to the bacterial chloramphenicol acetyl transferase (CAT) coding sequence following transfection to COS-1 cells. A model is proposed to account for the silencing of the protamine gene in all tissues except developing sperm cells.
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  • 86
    ISSN: 1573-5028
    Keywords: hypersensitive response ; Brassica campestris ; Xanthomonas campestris pv. vitians ; mRNA induction ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Xanthomonas campestris pv. vitians, a pathogen of lettuce, elicits a hypersensitive response within 12 hours of inoculation into Brassica leaves, characterized by tissue collapse, loss of membrane integrity, vein blockage and melanin production. In contrast, the compatible pathogen, X. c. pv. campestris, has no visible effects on leaves for 48 hours, after which inoculated areas show chlorosis which eventually spreads, followed by rotting. mRNA was prepared from leaves inoculated with suspensions of both pathovars or with sterile medium up to 24 hours following inoculation. In vitro translation of total and poly A+ RNA in rabbit reticulocyte lysate in the presence of 35S methionine followed by separation of the polypeptide products by 2D-PAGE, allowed comparison of the effects of these treatments on plant gene expression. Major changes in gene expression were observed as a consequence of the inoculation technique. In addition, after inoculation with X. c. vitians, up to fifteen additional major polypeptides appeared or greatly increased by four hours. Some of these had disappeared by nine hours and several more had appeared. No major polypeptides disappeared or decreased greatly in intensity following inoculation with X. c. vitians.
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  • 87
    ISSN: 1573-5028
    Keywords: coordinate regulation ; Cucurbita phytochrome cDNA ; gene expression ; light regulation ; multiple transcripts ; regulatory photoreceptor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have isolated several cDNA clones for phytochrome from a dicot, Cucurbita pepo L. cv. Black Beauty (zucchini), and have used them to study the regulation of Cucurbita phytochrome mRNA levels. A cDNA library was constructed from poly(A)+ RNA isolated from etiolated Cucurbita hypocotyl hooks and enriched for phytochrome mRNA by size fractionation. This library was screened with a 32P-labeled fragment isolated from an Avena phytochrome cDNA clone. Several putative phytochrome clones were isolated and mapped by restriction endonuclease analysis. On the basis of this analysis there is no evidence for the expression of multiple phytochrome genes in Cucurbita. Recent sequence analysis has confirmed that the largest of these clones, pFMD1 (∼3.6 kb), does indeed encode phytochrome and that it contains the entire amino acid coding sequence for Cucurbita phytochrome (33). RNA blot analysis has revealed that two polyadenylated phytochrome transcripts (∼5.6 kb and ∼4.2 kb) are present in both cotyledons and hypocotyl hooks of Cucurbita. In etiolated Cucurbita seedlings given a saturating pulse of red light, the abundance of both transcripts coordinately declines to 50–60% of the dark levels within 3 h and reaccumulates to dark levels within 24 h. Reversal of induction of this response by a far-red light pulse immediately following red light treatment is not observed, which is in contrast to the far-red reversibility of the red light promoted decrease in phytochrome mRNA abundance observed in Avena (6). Etiolated seedlings transferred to continuous white light also show a coordinate decrease in the levels of the two RNAs to ∼40% of the dark levels within 3 h. The magnitude of the light-induced decline in phytochrome mRNA abundance in Cucurbita is substantially less than the decrease previously reported for Avena (6).
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  • 88
    ISSN: 1573-5028
    Keywords: Bradyrhizobium ; gene expression ; nodulin ; soybean
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In vitro translation products of total RNA isolated from soybean nodules at successive stages of nodule development were analyzed by two-dimensional gel electrophoresis. In that way the occurrence of over 20 mRNAs specifically transcribed from nodulin genes was detected. The nodulin genes could be divided into two classes according to the time of expression during nodule development. Class A comprises at least 4 nodulin mRNAs which are found when a globular meristem is present in the root cortex. These class A nodulin genes have a transient expression. Class B nodulin genes are expressed when the formation of a nodule structure has been completed. Bradyrhizobium japonicum nod + fix-mutants, with large deletions spanning the nif H,DK region, still induced nodules showing normal expression of all nodulin genes, indicating that the nif H,DK region is not involved in the induction of nodulin genes. In nodules induced by Bradyrhizobium japonicum nod + fix-mutant HS124 the bacteria are rarely released from the infection thread and the few infected cells appear to be collapsed. All class A and class B nodulin genes are expressed in HS124 nodules with the exception of 5 class B genes.
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  • 89
    ISSN: 1573-5028
    Keywords: abscisic acid ; aleurone layer ; α-amylase ; gene expression ; gibberellic acid ; isozymes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The treatment of barley aleurone layers with gibberellic acid (GA3) results in the synthesis of two groups of α-amylase isozymes. Addition of abscisic acid (ABA) at the same time as GA3 inhibited the synthesis of both groups of isozymes. However, midcourse ABA addition (12 h or later after GA3) had a more inhibitory effect on the high pI α-amylase group than on the low pI α-amylase group. This midcourse inhibition was detectable within 2 h of ABA addition. Northern analysis results using cDNA probes for the high pI and low pI α-amylase groups paralleled the protein synthesis results for both isozyme groups. High pI α-amylase mRNA levels began to decrease within 2 h of midcourse ABA treatment and were less than 10% of the original level by 4 h. The levels of low pI α-amylase mRNA were decreased less by midcourse ABA addition than were high pI mRNA levels. Cordycepin and cycloheximide blocked the effects of midcourse ABA addition on α-amylase mRNA. These observations indicate that ABA inhibits α-amylase expression at the pretranslational level and that protein and RNA synthesis are required for midcourse ABA action to occur. Our results also show that α-amylase mRNA, which has been thought to be very stable, is degraded after midcourse ABA treatment.
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  • 90
    ISSN: 1573-5028
    Keywords: cucumber ; gene expression ; hydroxypyruvate reductase ; light regulation ; peroxisomal enzymes ; serine:glyoxylate aminotransferase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The development of peroxisomal enzymes in cotyledons of cucumber seedlings is strongly dependent on light. In light-grown seedlings, activities of two peroxisomal enzymes, hydroxypyruvate reductase (HPR) and serine: glyoxylate aminotransferase (SGAT), were barely detectable until three days postimbibition, after which time both activities increased rapidly and linearly for at least three days. In the dark, the activities of these enzymes increased slightly over the same time period, but only to about 5% to 10% of 7-day light-induced levels. When 51/2-day dark-grown seedlings were transferred into white light, activities of HPR and SGAT began to increase after approximately 8 h. HPR protein was shown by an immunoprecipitation assay to increase concurrently with enzymatic activity in both light- and dark-grown cotyledons. Immunoblotting results suggested that the amounts of SGAT-A and SGAT-B, the two subunits of SGAT, also developed along with SGAT activity. The relative levels of translatable mRNAs encoding HPR, SGAT-A, and SGAT-B were also light-dependent, and increased with a developmental pattern similar to enzyme activity and protein levels in light- and dark-grown cotyledons. In 51/2-day dark-grown cotyledons that were transferred to the light, translatable mRNAs for SGAT-A and SGAT-B began to increase within 1 h of illumination and continued of increase rapidly and linearly for the next 24 h in the light to a new steady-state level that was 45 times that of dark controls. Translatable HPR mRNA exhibited a biphasic pattern of accumulation, with a three-fold increase during the first 6 h of illumination, followed by an additional six-fold increase between 8 and 24 h. The accumulation of translationally active mRNA for both enzymes preceded the accumulation of the corresponding protein and enzyme activity by about 8 h. Our data suggest that the rise in enzyme activity depends on an increase in translatable mRNA for these enzymes and is regulated at a pretranslational level, most likely involving transcription of new mRNA.
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  • 91
    ISSN: 1573-5028
    Keywords: plant transformation ; CaMV promoters ; gene expression ; protein stability ; beta-conglycinin
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A gene encoding the α′-subunit of β-conglycinin was ligated to the 19S and 35S promoters of Cauliflower Mosaic Virus and introduced into petunia plants on a disarmed Ti-plasmid using Agrobacterium tumefaciens. Transformed cells were regenerated into whole plants and ummunoreactive polypeptides and hybridizable, polyadenylated mRNA were detected in transformed tissues. Expression from the 35S promoter was 10 to 50 times greater than expression from the 19S promoter. The level of immunodetectable polypeptides was greater in seeds than in leaves or callus tissue. In addition, the pattern of α′-polypeptide breakdown products was distinctive in seeds and leaves. We conclude that in seeds the higher levels of the α′-polypeptide reflect enhanced stability of this protein.
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  • 92
    Electronic Resource
    Electronic Resource
    Springer
    Plant cell, tissue and organ culture 8 (1987), S. 3-15 
    ISSN: 1573-5044
    Keywords: soybean ; plant transformation ; Agrobacterium ; Tiplasmid ; nopaline
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The response of Glycine max., G. soja and G. canescens genotypes to inoculation with different Agrobacterium strains was assessed. Percent visual tumor formation and tumor size varied widely among species and genotypes. Susceptible genotypes displayed a heightened response to nopaline strains of A. tumefaciens, relative to octopine, agropine, and A. rhizogenes strains. A nopaline strain engineered to contain a chimeric neomycin phosphotransferase II gene conferred kanamycin resistance to soybean tissue at kanamycine levels as high as 300 μg/ml.
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  • 93
    Electronic Resource
    Electronic Resource
    Springer
    Molecular and cellular biochemistry 75 (1987), S. 61-70 
    ISSN: 1573-4919
    Keywords: gene expression ; development ; tissue-specific expression ; cell cycle-dependent genes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract 4F1, 2A9 and 2F1 represent three of a number of cDNA sequences which have been identified because their cognate RNAs markedly increase when quiescent cells in culture are stimulated with serum. Studies using a variety of cell culture systems have shown that the expression of these genes is modulated by various growth factors and mitogens and thus such genes are considered to be ‘growth-associated.’ Thus far, little information has been obtained with these in vitro systems about the function of these genes. In an attempt to begin to elucidate the role of these genes (if any) in the physiology of the normal cell, we have analyzed the levels of 4F1, 2A9 and 2F1 transcripts in a variety of differentiated organs and tissues of adult and fetal rats. Our results show that each of these growth-associated genes exhibits its own unique pattern of expression, unrelated to the proliferative activity of the tissue. These data suggest that these genes most likely do have specific functions in normal tissue in addition to their role in the induction of DNA synthesis in quiescent cells in culture.
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  • 94
    Electronic Resource
    Electronic Resource
    Springer
    Molecular and cellular biochemistry 78 (1987), S. 3-8 
    ISSN: 1573-4919
    Keywords: polyamines ; RNA polymerase ; transcription ; gene expression ; mycobacteria
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract All three polyamines — putrescine, spermidine and spermine stimulated the activity of mycobacterial RNA polymerase in vitro although the concentration required for maximal stimulation was different for each of the amines. Spermidine and spermine showed a biphasic effect on the enzyme activity. Stimulation of RNA synthesis by spermidine occurs only at higher DNA template/enzyme ratio. Spermidine stimulates RNA synthesis by acting on the elongation phase of RNA synthesis but it had no effect on initiation phase. Addition of mycobacterial RNA to the assay mixture resulted in the inhibition of RNA polymerase activity and this inhibition could be reversed by spermidine suggesting that spermidine stimulates transcription by binding to nascent RNA and thus destabilizing the short DNA-RNA hybrid region.
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  • 95
    Electronic Resource
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    Springer
    Molecular genetics and genomics 209 (1987), S. 475-480 
    ISSN: 1617-4623
    Keywords: Agrobacterium ; Hairy root ; T-DNA ; Transgenic tobacco
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Segments of the TL-DNA of the agropine type Ri plasmid pRi 1855 encompassing single and groups of open-reading frames were cloned in the Ti plasmid-derived binary vector system Bin 19. Leaf disc infections on Nicotiana tabacum led to transformed plants, some of which showed typical hairy root phenotypes, such as the wrinkled leaf morphology, excessive and partially non geotropic root systems and the ability of leaf explants to differentiate roots in a hormone-free culture medium. Particularly interestingly, most of these traits were shown by plants transformed with a TL-DNA segment encompassing the single ORF 11, corresponding to the rolB locus. Hairy root can be induced by this latter T-DNA segment on wounded stems of tobacco plants; hairy root induction on carrot discs requires, on the contrary, a more complex complement of TL-DNA genes.
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  • 96
    ISSN: 1617-4623
    Keywords: Agrobacterium ; Ti plasmids ; Cytokinin biosynthesis ; Isopentenyltransferase ; trans-zeatin
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Tzs and ipt are two Ti plasmid genes coding for proteins with isopentenyltransferase (IPT) activity in vitro. We cloned both genes for protein expression in Escherichia coli and in Agrobacterium tumefaciens, and we investigated differences between the two genes by analysing the properties of the proteins in vitro and in vivo. In vitro, extracts with tzs or ipt-coded proteins had high IPT activity, and the enzymes were identical in most properties. The most important difference was detected in vivo: the tzs-encoded protein was very active in cytokinin production, while the ipt protein required overexpression in order to obtain measurable activity in bacteria. In both cases, rans-zeatin was the major product of the gene activity. Formation of this cytokinin requires a hydroxylase function in addition to the IPT reaction. No such activity could be ascribed to tzs or ipt-encoded proteins in vitro or in vivo, but cytokinin hydroxylase activity was detected in cells and extracts of E. coli, regardless of the presence or absence of the cytokinin genes. Based on these results it is proposed that both genes code for a single enzyme activity (isopentenyltransferase), that the genes and proteins are adapted for function either in bacteria (tzs) or in transformed plant cells (ipt), and that in both prokaryotic and eukaryotic cells hydroxylation to trans-zeatin is a function contributed by host enzymes.
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  • 97
    Electronic Resource
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    Springer
    Molecular genetics and genomics 206 (1987), S. 1-8 
    ISSN: 1617-4623
    Keywords: Ri plasmid ; Agrobacterium ; Mini replicon ; Replication origin ; Incompatibility
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Starting from a cosmid library of the 250 kb hairy root inducing plasmid pRiA 4b, a mini-pRiA 4b replicon of 4.6 kb was isolated. This mini-plasmid was stably maintained in Agrobacterium species and its replication characteristics, such as temperature-sensitive replication, copy number and incompatibility, were similar to those of the parental pRiA 4b. Analysis of deletion mutants indicated that almost the entire 4.6 kb region was required for autonomous replication. The nucleotide sequence of mini-pRiA 4b was determined by the chain-termination method. Three large open reading frames, which are likely to contribute to the replication of pRiA 4b, were identified in the same orientation along the sequence.
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  • 98
    Electronic Resource
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    Springer
    Molecular genetics and genomics 206 (1987), S. 192-199 
    ISSN: 1617-4623
    Keywords: Poplar ; Transformation ; Agrobacterium
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A plant transformation and regeneration system has been developed for Populus species. Leaf explants, from stabilized shoot cultures of a Populus hybrid NC-5339 (Populus alba x grandidentata), were co-cultivated with Agrobacterium tumefaciens on a tobacco nurse culture. Both oncogenic and disarmed strains of A. tumefaciens harboring a binary vector which contained two neomycin phophotransferase II (NPT II′) and one bacterial 5-enolpyruvylshikimate 3-phosphate (EPSP) synthase (aroA) chimeric gene fusions were used. Shoots did not develop when leaf explants were co-cultivated with the binary disarmed strain of A. tumefaciens. However, transformed plants with and without the wild type T-DNA were obtained using an oncogenic binary strain of A. tumefaciens. Successful genetic transformation was confirmed by NPT II′ enzyme activity assays, Southern blot analysis and immunological detection of bacterial EPSP synthase by Western blotting. This is the first report of a successful recovery of transformed plants of a forest tree and also the first record of insertion and expression of a foreign gene of agronomic importance into a woody plant species.
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  • 99
    ISSN: 1617-4623
    Keywords: Nicotiana tabacum ; Agrobacterium ; Plant transformation ; Genetic stability ; Kanamycin resistance
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Two lines of transgenic Nicotiana tabacum transformed to kanamycin resistance by means of a binary Agrobacterium vector containing a nos-npt gene were investigated over three generations. Southern hybridization and crossing analyses revealed that a single copy of T-DNA had integrated in each line and that the kanamycin resistance was regularly transmitted to the progeny as a monogenic dominant trait. Homozygous transgenic plants were fully fertile, morphologically normal and did not significantly differ from wild-type plants in the quantitative characters examined (plant height, flowering time). The two lines showed very low, but significantly different levels of meiotic instability: kanamycin-sensitive plants occurred among backcross progeny from homozygous transgenic plants with frequencies of 6/45,000 and 25/45,000, respectively. The sensitive plants arose independently of each other and thus resulted from meiotic rather than mitotic events. These findings demonstrate for the first time that integrated foreign genes can be transmitted to progeny with the high degree of meiotic stability required for commercial varieties of crop plants. They emphasize the importance of non-homologous integration and of avoiding co-integration of inactive gene copies for achieving meiotically stable transformants.
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  • 100
    ISSN: 1617-4623
    Keywords: Agrobacterium ; Monocotyledonous plants ; Plant factors ; T-DNA circularization ; vir gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary T-DNA circularization is one of the molecular events specifically induced in agrobacterial cells upon their infection of dicotyledonous plant cells. We developed a seedling co-cultivation procedure to determine whether or not monocotyledonous plants have the ability to induce T-DNA circularization and vir gene expression. Co-cultivation of Agrobacterium tumefaciens with seedlings of dicotyledonous plants showed that the circularization event takes place efficiently. The exudates and extracts of the seedlings also effectively induced T-DNA circularization and vir gene expression, indicating that dicotyledonous seedlings contain diffusible factors capable of inducing these molecular events. In contrast, neither T-DNA circularization nor vir gene expression was detectable when Agrobacterium was incubated with seedlings of monocotyledonous plants. Supplementing with acetosyringone, a known inducer of vir gene expression and T-DNA circularization, resulted in the induction of circularization during co-cultivation with monocotyledonous seedlings. These results indicate that the seedlings of monocotyledonous plants have no detectable amounts of diffusible inducers, unlike dicotyledonous seedlings. Therefore, it is unlikely that the vir genes are expressed in Agrobacterium inoculated in monocotyledonous plants. This may be one of the blocks in tumorigenesis of monocotyledonous plants by Agrobacterium.
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