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  • 1985-1989  (105)
  • 1955-1959  (7)
  • 1950-1954
  • 1940-1944
  • Agrobacterium
  • Brassica napus
  • 1
    Electronic Resource
    Electronic Resource
    Springer
    Sexual plant reproduction 2 (1989), S. 15-17 
    ISSN: 1432-2145
    Keywords: Brassica napus ; Spontaneous androgenesis ; Erucic acid content ; Flower colour
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Seeds of androgenetic origin were obtained among the F1 progenies of two crosses between resynthesized and cultivated forms of Brassica napus. The high-erucic, white-flowered, resynthesized line No7076 acted as the female, and the zero-erucic, yellow-flowered, cultivars ‘Topas’ and ‘Puma’, as males. No androgenetic seeds were obtained in the reciprocal crosses. Resynthesized rape could thus be of potential use for the production of androgenetic plants. Of special interest is the high frequency (21%) of spontaneous androgenesis observed in one of the two crosses. One plant, determined from erucic acid content and flower colour analysis as androgenetic, had a diploid chromosome number. Further knowledge about the genetic control of spontaneous androgenesis in the present material and the origin of the cytoplasm in androgenetic plants are required in order to exploit this phenomenon in practical plant breeding.
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  • 2
    Electronic Resource
    Electronic Resource
    Springer
    Plant cell reports 8 (1989), S. 303-306 
    ISSN: 1432-203X
    Keywords: Ti plasmid ; Virulence ; Brassica napus ; Brassica juncea
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Brassica napus and Brassica juncea were infected with a number of Agrobacterium tumefaciens strains. Tumourigenesis was very rapid and extremely efficient on B. juncea with all but one of the strains. Tumourigenesis on B. napus varied widely. It was very efficient with the nopaline strains, was reduced with the succinamopine strain A281 and was very weak with the octopine strains. The latter observation was confirmed with six different B. napus rapeseed cultivars. The selectivity was due to differences in the virulence of Ti plasmids with B. napus, rather than the tumourigenicity of the T-DNA or virulence of the chromosomal genes associated with the strains. An exception was strain LBA4404. The virulence of the octopine strains was increased by coinfection with more virulent disarmed strains and by induction with acetosyringone.
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  • 3
    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.
    Type of Medium: Electronic Resource
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  • 4
    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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  • 5
    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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  • 6
    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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  • 7
    Electronic Resource
    Electronic Resource
    Springer
    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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  • 8
    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.
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  • 9
    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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  • 10
    Electronic Resource
    Electronic Resource
    Springer
    Theoretical and applied genetics 77 (1989), S. 489-494 
    ISSN: 1432-2242
    Keywords: Microspore culture ; Spontaneous diploids ; Erucic acid ; Inheritance ; Brassica napus
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The levels of erucic acid and other fatty acids in seeds of microspore-derived spontaneous diploid plants from crosses between low and high erucic acid parents were examined. The analysis confirmed that erucic acid is simply inherited and is determined by two genes that act in an additive manner. The effects of the genes for erucic acid on the levels of the other fatty acids was also determined and many significant correlations were found. In particular, erucic acid levels were negatively correlated with oleic acid and linoleic acid levels. The study also illustrates several advantages of using haploidy to analyze the inheritance of agronomically important traits. In particular, the number of phenotypic classes is smaller in androgenic populations and differences between classes are greater than in an F2 population.
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  • 11
    Electronic Resource
    Electronic Resource
    Springer
    Theoretical and applied genetics 77 (1989), S. 651-656 
    ISSN: 1432-2242
    Keywords: Brassica napus ; Brassica nigra ; Somatic hybrids ; Resistance to Phoma lingam
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Brassica napus and B. nigra were combined via protoplast fusion into the novel hybrid Brassica naponigra. The heterokaryons were identified by fluorescent markers and selected by flow sorting. Thirty hybrid plants were confirmed by isozyme analysis to contain both B. nigra and B. napus chromosomes; of these, 20 plants had the sum of the parental chromosome numbers. A non-random segregation of the chloroplasts was found in the hybrids. Of 14 hybrid plants investigated, all had the B. napus type of chloroplast. The resistance to Phoma lingam found in the B. nigra cultivar used in the fusion experiments was expressed in 26 of the hybrid plants. The hybrids obtained in this study contain all of the three Brassica genomes (A, B and C) and have thus created unique possibilities for genetic exchanges between the genomes. Since most of the plants were fertile as well as resistant to P. lingam, they have been incorporated into conventional rapeseed breeding programs.
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  • 12
    Electronic Resource
    Electronic Resource
    Springer
    Theoretical and applied genetics 77 (1989), S. 721-728 
    ISSN: 1432-2242
    Keywords: Brassica napus ; Introgression ; Adaptation to short day ; Photo-insensitivity ; Oligogenic control
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Among the oleiferous Brassicas, B. napus has the highest seed and oil productivity. As it is a species adapted to the temperate regions, its spring type is either unable to flower or flowers too late in the short — day winter (rabi) season of the subtropics. B. napus (genome AACC) is an amphidiploid between B. campestris (AA) and B. oleracea (CC), and shares one genome with the other allotetraploids B. juncea (AABB) and B. carinata (BBCC). While B. napus lacks ecotypes adapted to the subtropics, the other four species are well represented in this climatic zone. Reciprocal crosses with or without one direct backcross to B. napus have been carried out with the intention of transfering short-day adaptability. The aim was to introgress the A genome of carefully selected early representatives of B. campestris and B. juncea with the corresponding genome in B. napus, and similary the C genome from B. oleracea and B. carinata with the analogous genome in B. napus. B. campestris, B. juncea and the clearly later species, B. oleracea var ‘alboglabra’ and B. carinata, seem to be almost equally effective in introgressing the appropriate earliness necessary for growth in Bangladesh. One backcross sligthly delayed segregation of early types. Convergent crosses did not result in the transgression of earliness, which was unexpected since the inheritance of flowering and maturity indicated a polygenic regulation. This result is partly explained by assuming dominant oligogenic control of the photoperiodic response. Introgression of earliness with the C genome doesn't seem to be necessarily related with the earliness of the donor species. Intergenomic interactions may be important. Interesting new lines were selected with high yield. Thus there is a good probability that Bangladesh will have a new oil crop. As these lines were observed to be early in Sweden as well, they could potentially push rapseed cultivation further north in temperate regions where the growing period is limited by short summers.
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  • 13
    ISSN: 1432-2242
    Keywords: Agrobacterium-transformation ; Asymmetric protoplast fusion ; Brassica napus ; Brassica nigra
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary With the idea to develop a selection system for asymmetric somatic hybrids between oilseed rape (Brassica napus) and black mustard (B. nigra), the marker gene hygromycin resistance was introduced in this last species by protoplast transformation with the disarmed Agrobacterium tumefaciens strain C58 pGV 3850 HPT. The B. nigra lines used for transformation had been previously selected for resistance to two important rape pathogens (Phoma lingam, Plasmodiophora brassicae). Asymmetric somatic hybrids were obtained through fusion of X-ray irradiated (mitotically inactivated) B. nigra protoplasts from transformed lines as donor with intact protoplasts of B. napus, using the hygromycin resistance as selection marker for fusion products. The somatic hybrids hitherto obtained expressed both hygromycin phosphotransferase and nopaline synthase genes. Previous experience with other plant species had demonstrated that besides the T-DNA, other genes of the donor genome can be co-transferred. In this way, the produced hybrids constitute a valuable material for studying the possibility to transfer agronomically relevant characters — in our case, diseases resistances — through asymmetric protoplast fusion.
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  • 14
    ISSN: 1432-2242
    Keywords: Brassica napus ; Protoplast fusion ; Triazine resistance ; Cytoplasmic male sterility ; Mitochondrial DNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Fusion of leaf protoplasts from an inbred line of Brassica oleracea ssp. botrytis (cauliflower, n=9) carrying the Ogura (R1) male sterile cytoplasm with hypocotyl protoplasts of B. campestris ssp. oleifera (cv “Candle”, n=10) carrying an atrazine-resistant (ATR) cytoplasm resulted in the production of synthetic B. napus (n=19). Thirty-four somatic hybrids were produced; they were characterized for morphology, phosphoglucose isomerase isoenzymes, ribosomal DNA hybridization patterns, chromosome numbers, and organelle composition. All somatic hybrids carried atrazine-resistant chloroplasts derived from B. campestris. The mitochondrial genomes in 19 hybrids were examined by restriction endonuclease and Southern blot analyses. Twelve of the 19 hybrids contained mitochondria showing novel DNA restriction patterns; of these 12 hybrids, 5 were male sterile and 7 were male fertile. The remaining hybrids contained mitochondrial DNA that was identical to that of the ATR parent and all were male fertile.
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  • 15
    Electronic Resource
    Electronic Resource
    Springer
    Theoretical and applied genetics 78 (1989), S. 473-479 
    ISSN: 1432-2242
    Keywords: Brassica napus ; Microspore and anther culture comparison ; Androgenic lines ; Days to flower ; Days to maturity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A direct comparison of microspore culture and anther culture was made in Brassica napus using F1 crosses of Regent (canola) by Golden (rapeseed), and their reciprocals, as well as a hybrid between Reston and a highly embryogenic, canola-quality breeding line (G231) as donor plants. The study confirmed that microspore culture can be ten times more efficient than anther culture for embryo production. Embryo yields from cultures initiated from the Reston x G231 were four-fold greater than those initiated from the Regent x Golden crosses, and significant differences were also detected among cultures initiated from the different Regent x Golden crosses. These results illustrate the influence that donor plant genotype has on embryo production. However, superior embryogenic potential among donor material was not always coincident with superior plant production. The average haploid-todiploid ratio in microspore-derived regenerates was 2∶1 for the population obtained from the Regent x Golden crosses but 1∶1 for the Reston x G231 cross. For both types of material, the frequency of diploids increased upon repeated cycles of explanting. A field study showed that there were no differences between the populations of anther-derived and microspore-derived spontaneous diploid and doubled haploid lines, with respect to the days required for them to flower or to mature. The information is valuable for canola breeding programs considering the use of haploidy.
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  • 16
    Electronic Resource
    Electronic Resource
    Springer
    Theoretical and applied genetics 78 (1989), S. 161-168 
    ISSN: 1432-2242
    Keywords: Brassica napus ; Transgenic ; Heavy metal tolerance ; Human metallothionein gene ; Ti-plasmid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A chimeric gene containing a cloned human metallothionein-II (MT-II) processed gene was introduced into Brassica napus and Nicotiana tabacum cells on a disarmed Ti-plasmid of Agrobacterium tumefaciens. Transformants expressed MT protein as a Mendelian trait and in a constitutive manner. Seeds from self-fertilized transgenic plants were germinated on media containing toxic levels of cadmium and scored for tolerance/ susceptibility to this heavy metal. The growth of root and shoot of transformed seedlings was unaffected by up to 100 μM CdCl2, whereas control seedlings showed severe inhibition of root and shoot growth and chlorosis of leaves. The results of these experiments indicate that agriculturally important plants such as B. napus can be genetically engineered for heavy metal tolerance/sequestration and eventually for partitioning of heavy metals in non-consumed plant tissues.
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  • 17
    Electronic Resource
    Electronic Resource
    Springer
    Theoretical and applied genetics 78 (1989), S. 793-797 
    ISSN: 1432-2242
    Keywords: Brassica napus ; Rapeseed ; Fatty acid composition ; Linolenic acid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Results from a diallel mating of two rapeseed lines with distinctly different linolenic acid concentration show that this trait is mainly under control of nuclear genes of the embryo. However, significant differences in reciprocal F1, BC1 and BC2 indicate maternal control, which is realized by interaction between maternal genotype and nuclear genes of the embryo. Additionally, temperature exerts considerable influence on the degree of maternal control. Since no reciprocal differences are detectable in F2, cytoplasmic factors seem not to be involved in the inheritance of linolenic acid concentration. Hypotheses on the physiological nature of maternal control of this trait are discussed.
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  • 18
    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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  • 19
    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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  • 20
    ISSN: 1615-6102
    Keywords: Brassica napus ; Vegetative cell ; Microbodies ; Pollen
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The ultrastructure of the vegetative cell ofBrassica napus tricellular pollen grains, just before anthesis with standard chemical fixation, is reported. The vegetative cell may be regarded as a highly differentiated and metabolically active fat-storage cell. It contains many mitochondria with a well developed internal membrane system, starchless plastids, microbodies, lipid bodies, dictyosomes and numerous vesicles thought to originate from the dictysomes. Rough endoplasmic reticulum organized in stacks of cisternae is also spatially associated with certain organelles, mainly lipid bodies, microbodies and plastids. There are also randomly distributed polyribosome areas. The microbodies are mainly polymorphic in shape and are often observed in contact with lipid bodies. The above spatial relationship implies that the microbodies may have a glyoxysomal function. In the late period of vegetative cell maturation, the microbodies are probably involved in the process of glyconeogenesis in which the conversion of lipid reserves to sugar takes place.
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  • 21
    ISSN: 1615-6102
    Keywords: Brassica napus ; Cell division ; Male germ unit ; Pollen ; Sperm cells
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The association of the two sperm cells inBrassica napus pollen following the generative cell division was investigated. The generative cell during division is located in the center of the pollen grain, within the vegetative cell. The space present between the two cells is slightly irregular as seen following standard glutaraldehyde fixation. After completion of mitosis vesicles appear in the equatorial plane, coalescing centripetally to form a cell plate which fuses with the membrane of the generative cell, dividing it in two sperm cells. They are isolated from the vegetative cell by the space between the two cell membranes and are separated from each other by a similar space resulting from the cell plate formed during cytokinesis.
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  • 22
    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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  • 23
    ISSN: 1617-4623
    Keywords: Acetolactate synthase ; DNA sequence ; Brassica napus ; Oilseed rape ; Herbicide resistance
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary An acetolactate synthase gene was isolated and characterized from Brassica napus. This B. napus acetolactate synthase gene encodes a deduced polypeptide sequence of 637 amino acids which is 85% homologous to the corresponding proposed gene product from Arabidopsis thaliana. Peptide domains recently associated with herbicide resistance/sensitivity are conserved between the two sequences. From Southern analysis we conclude that the gene isolated is one member of a multigene acetolactate synthase gene family comprising four or five members. A probe spanning the presumptive transit peptide sequence of this gene was shown by Southern analysis to hybridize to a unique sequence in the B. napus genome. This unique probe was used to analyse DNA from B. campestris and B. oleracea, the presumed progenitors of B. napus. On the basis of restriction fragment length polymorphism, we conclude that the B. napus gene isolated here originated in B. campestris. Total acetolactate synthase-homologous transcripts were analysed in a variety of B. napus tissues, and showed preferential accumulation in rapidly growing material. The genomic clone was mutated in vitro at codon 173 to replace a proline residue with serine. This was re-introduced into plants, using Agrobacterium vectors, producing a herbicide-resistant phenotype which is characteristic of the predicted gene product.
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  • 24
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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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  • 25
    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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  • 26
    ISSN: 1573-5060
    Keywords: Diplotaxis erucoides ; Brassica napus ; rapeseed ; ovary culture ; intergeneric hybridization ; cytogenetics ; addition lines ; cytoplasmic male sterility
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Intergeneric hybrids (F1) Diplotaxis erucoides (DeDe) x Brassica napus (AACC) and the first backcross to B. napus (BC1) have been obtained through in vitro culture of excised ovaries. The chromosome numbers of F1 and BC1 plants proved the occurrence of unreduced gametes. The study of metaphase I chromosome pairing showed that autosyndesis in De genome and allosyndesis between De and A/C genomes might exist. The male fertility of the F1 plants was low. Some male-sterile plants were found in F1 and BC1 progeny. The possibilities of creating addition lines B. napus-D. erucoides and of obtaining a new cytoplasmic male sterility in B. napus are discussed.
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  • 27
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    Euphytica 42 (1989), S. 135-140 
    ISSN: 1573-5060
    Keywords: Brassica napus ; swede breeding ; inter-plot competition ; dry-matter yield
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Two experiments were done to evaluate the use of single-row plots for assessing dry-matter yield of swedes. In the first experiment the correlation between the dry-matter yields of 20 cultivars in unguarded single-row plots and those in guarded double-row plots was poor (r=0.39). It was improved (r=0.58) by adjusting the single-row plot fresh-weight yields for inter-plot competition, through an analysis of covariance in which the covariate was the mean of the adjacent plot yields, and then estimating the pure stand dry-matter yields. However, the correlation was still lower than that between unguarded double-row plots and the guarded double-row plots (r=0.73), and this latter correlation was also improved (r=0.82) by adjusting the fresh-weight yields of the unguarded plots for inter-plot competition. The second experiment provided an explanation for the partial success of the covariance adjustment. The individual neighbour effects of six cultivars were determined. Doon Major and Ruta Øtofte decreased the yields of adjacent cultivars whereas Dryden and Melfort increased them. Angela and Magres had little effect. The correlation between the dry-matter yield neighbour effects of cultivars and their fresh-weight yields was high (r=−0.81), but not complete. So although competitive ability was associated with root fresh-weight yield, other factors must also be involved. It was concluded that a two-replicate trial with unguarded double-row plots is preferable to a four-replicate trial with unguarded single-row plots for dry-matter yield assessment of early generation families in a pedigree swede breeding programme.
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  • 28
    ISSN: 1573-5060
    Keywords: Brassica napus ; forage rape ; swedes ; self-incompatibility ; S-genes ; hybrid breeding
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Seed set after self-pollination confirmed that 19 lines of Brassica napus were self-incompatible. Eight lines, H, J, Q, W, X, K, P and Z, were fully cross-compatible. Line R was cross-compatible with these lines but often had a low seed set as female parent. These results are consistent with the activity of nine distinct S-alleles. Line S was cross-incompatible with K, as was V with P, and F with Z. With each of the lines A, E, B, O, G, L and M at least one reciprocal difference was found so that the number of additional distinct S-alleles could not be inferred, but there must be a minimum of seven. Pollen tube counts of intra- and inter-line pollinations using M, B and X confirmed the homozygosity of these lines with respect to self-incompatibility status and the presence of non-reciprocal compatibility between M and B. The results are interpreted in terms of the activity of both B. oleracea and B. campestris S-loci and the implications for hybrid breeding are discussed.
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  • 29
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    Euphytica 42 (1989), S. 99-103 
    ISSN: 1573-5060
    Keywords: Brassica napus ; self-incompatibility
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary The interactions of nine S-gene lines of Brassica napus were examined. Seven different S-genes were involved in the crosses, with three lines of different origin having the same S-gene. In general, the reactions on selfing the heterozygotes were as expected from the results of test crosses with the homozygous parent lines. All types of dominance relationships were found, including reversal of dominance in pollen and pistil. Weakening of activity of one or both genes was found in several cases. Background genotype was found have to some effect and reciprocal differences in reactions were also observed.
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  • 30
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    Plant cell, tissue and organ culture 17 (1989), S. 153-158 
    ISSN: 1573-5044
    Keywords: Brassica napus ; spring cultivars ; protoplast culture ; plant regeneration ; cultivar variation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Protoplast cultures were prepared from hypocotyls of ten spring rapeseed cultivars. Protoplasts from all genotypes tested formed calli, and shoots were regenerated from calli of nine of the genotypes at frequencies varying from 15 to 76%. The regenerating cultivars fell into a high regenerating group (〉60% and a low regenerating group 〈25%).
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  • 31
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    Plant cell, tissue and organ culture 18 (1989), S. 129-141 
    ISSN: 1573-5044
    Keywords: Brassica napus ; organogenesis ; plantlet formation ; regeneration ; root segments ; shoot bud differentiation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Root segments obtained from aseptically germinated seedlings of Brassica napus cv. Westar were used to optimize conditions for high-frequency shoot bud differentiation. The presence of low kinetin (0.5 μM) and relatively high indole-butyric acid (1.0 μM) levels facilitated optimum shoot bud differentiation. Modified MS medium (MMS) was superior to the other three basal media tested (MS, B5 and White's). Elevated sodium dihydrogen phosphate levels increased the differentiation of shoot buds. Increasing or decreasing the level of sucrose from 3% reduced the frequency of explants forming shoot buds. Addition of glutamine enhanced both the frequency of responding explants, as well as the number of shoots per responding explant. Root segments from 13-day-old seedlings produced the highest response (58%) in the presence of 100 mg l-1 glutamine. The position of the segment on the main root, size, and the presence or absence of lateral roots altered the morphogenic response. Sealing of the donor seedling cultures with Parafilm® instead of Stretch' n seal® resulted in a higher production of shoot buds, although root segment cultures were not affected by the type of sealing. Spontaneous rooting occurred on all developed shoots.
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  • 32
    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.
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  • 33
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    Methods in cell science 12 (1989), S. 139-144 
    ISSN: 1573-0603
    Keywords: microinjection ; genetic transformation ; protoplasts ; microspores ; Brassica napus
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary This paper describes a general method suitable for the microinjection ofBrassica napus protoplasts, unicellular microspores, and multicellular microspores. By incorporating components taken from other methods, manual operations frequently involved in the microinjection of plant cells have been simplified and microinjection rates increased. The embedding of cells in agarose provides a simple alternative to the variety of sophisticated immobilization strategies devised for different plant cell types thereby reducing the manipulations often involved in the culture of microinjected cells. Use of an automatic microinjector eliminated the operation of fine control systems on manual injectors; however, precision in sample delivery was reduced. Analyses indicate that transformed tissues can be recovered from microinjected protoplasts and microspores at high frequencies.
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  • 34
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    Plant and soil 113 (1989), S. 155-160 
    ISSN: 1573-5036
    Keywords: Brassica napus ; Ca uptake ; cation-anion balance ; rock phosphate ; split pot experiments
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Rape and sunflower were compared with respect to their rock phosphate mobilizing capacities, cationanion balance and uptake of Ca and NO3 at P-starvation. Rape was able to mobilize P from rock phosphate, whereas sunflower was not. When grown on a complete nutrient solution with NO3 as the only nitrogen source, both species took up more nutrient anions than cations. Withholding phosphate from the nutrient solution did not change the uptake pattern of rape, but sunflower took up more nutrient cations than anions at P-starvation, due to a strong decline in NO3 uptake. With both species, Ca uptake was not affected by phosphate in the nutrient solution. In split pot experiments, with rock phosphate supplied spatially separated from other nutrients, rape was still able to mobilize rock phosphate. A high Ca concentration had no effect on this capacity. The results indicated that in our experiments rock phosphate mobilization by rape was not associated with an excess of cation over anion uptake and neither with a high Ca uptake rate.
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  • 35
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    Plant and soil 113 (1989), S. 161-165 
    ISSN: 1573-5036
    Keywords: agar plate technique ; Brassica napus ; citric acid ; malic acid ; phosphate nutrition ; rhizosphere acidification
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Local rhizosphere acidification by rape as a reaction to P-starvation was visualized by means of an agar plate technique. By means of a modification of this technique local differences in cation-anion uptake and organic acid exudation along intact roots of rape were observed for plants grown on nutrient solution with or without added P. No differences in uptake rates of K-, NO3- and Ca-ions could be detected between P-starved and P-supplied plants. However, exudation of malic and citric acid was distinctly higher in acidified root zones of P-starved plants, coinciding with higher levels of malate in the corresponding root tissue. Organic acid exudation is indicates as the cause of local rhizosphere acidification by rape as a reaction to P-starvation and as a possible mechanism of its phosphate-solubilizing capacity.
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  • 36
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    Plant cell, tissue and organ culture 19 (1989), S. 213-224 
    ISSN: 1573-5044
    Keywords: Brassica napus ; B. oleracea ; rapid-cycling brassica populations ; protoplasts ; regeneration ; maltose
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Protoplasts were isolated from aseptic shoot cultures of commercial cultivars ofBrassica napus, B. oleracea andB. campestris, and from the six ‘rapid-cycling brassica species’. Of the rapid-cycling species, onlyB. napus responded well to the culture conditions used; 2% of protoplasts formed calli and up to 5% of calli regenerated shoots. Regeneration was also achieved from commercial cultivars ofB. napus andB. oleracea. For these two species the plating density, time of dilution with fresh medium and the composition of the shoot-inducing medium were all found to have an important influence on the efficiency of plant regeneration. Both responded better to maltose than to sucrose-based media. Under the optimum conditionsB. napus showed a plating efficiency of 7.8% and shooting efficiency of 17%; forB. oleracea the figures were 2% and 56%, respectively.
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  • 37
    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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  • 38
    ISSN: 1432-2145
    Keywords: Brassica napus ; Cytoplasmic male sterility ; Polypeptides ; Temperature ; Restoration
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Polypeptides were extracted from stamens of a rapeseed (Brassica napus) cultivar, Regent, a near isogenic male-sterile line, Polima-R7 (Pol-R7), and a high-temperature-restored malefertile Pol-R7 (TR) and subsequently separated by two-dimensional isoelectric focusing-polyacrylamide gel electrophoresis under denaturing conditions. Four variable polypeptides with a pI around 6 were observed. Two stamen polypeptides (40000 Da, 38000 Da) were unique to Regent, and the other two (32000 Da, 30000 Da) were unique to the male-sterile Pol-R7. When the male-sterile Pol-R7 was treated with day/night temperatures of 30°/24° C for 7–10 days prior to flowering, both polypeptides unique to Regent reappeared, while the smaller polypeptides disappeared. Temperature-restored male-fertile Pol-R7 (TR) produced fertile pollen, while its short stamen filaments resembled those of the male-sterile Pol-R7. These changes in protein expression may be causally related to the CMS phenotype.
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  • 39
    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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  • 40
    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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  • 41
    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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  • 42
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    Theoretical and applied genetics 75 (1988), S. 438-445 
    ISSN: 1432-2242
    Keywords: Brassica napus ; Rapeseed ; Transformation ; Agrobacterium tumefaciens ; Ti plasmid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Agrobacterium-mediated transformation of thin cell layer explants (Klimaszewska and Keller 1985) yielded large numbers of transgenic plants of a major Canadian rapeseed cultivar Brassica napus ssp. oleifera cv Westar. The morphology and fertility of these plants were indistinguishable from controls. The Ti plasmid vector, pGV3850 (Zambryski et al. 1983) was used as a cis vector and as a helper plasmid for the binary vector pBin19 (Bevan 1984). Selectable marker genes that conferred resistance to high levels of kanamycin (Km) on Nicotiana tabacum were less efficient in the selection of transgenic B. napus. At low levels of Km (15 μg/ml) large numbers of transgenic plants (50%) were identified among the regenerants by nopaline synthase activity and several of these were confirmed by Southern blot analyses. Only a small number were resistant to higher levels of Km (80 μg/ml). Preliminary analyses indicated that resistance to Km was transmitted to the selfed progeny. Chimeric chloramphenicol acetyl transferase genes were ineffective biochemical markers in transgenic B. napus.
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  • 43
    ISSN: 1432-2242
    Keywords: Brassica napus ; Oilseed rape ; Genetransfer ; Seed storage protein gene ; Tissue-specific expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Genetically transformed plants of Brassica napus L. (oilseed rape) were obtained from hypocotyl expiants using Agrobacterium tumefaciens vectors. Hypocotyl explants were inoculated with disarmed or oncogenic A. tumefaciens strains, EHA101 and A281, and then cultured on media containing kanamycin. The A. tumefaciens strains harbored a binary vector, which contained a neomycin phosphotransferase II (NPTII) gene driven by the 35S promoter of cauliflower mosaic virus and an engineered napin (seed storage protein) gene with its own promoter (300 nucleotides 5′ to the start of translation). Transformation of B. napus plants was confirmed by detection of NPT II enzyme activity, Southern blot analysis and inheritance of the kanamycin-resistance trait (NPT II gene) in the progeny. Expression of the engineered napin gene in embryos but not in leaves of transgenic plants was observed by Northern analysis. These data demonstrate that morphologically normal, fertile transgenic B. napus plants can be obtained using Agrobacterium as a gene vector and that developmentally regulated expression of reintroduced genes can be achieved.
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  • 44
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    Theoretical and applied genetics 75 (1988), S. 546-552 
    ISSN: 1432-2242
    Keywords: Brassica napus ; Brassica hirta ; Somatic hybridization ; mtDNA ; ctDNA ; rDNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Somatic hybridization between Brassica napus and B. hirta (or Sinapis alba) is described. No cybrid plant with B. napus nucleus exhibiting cytoplasmic male sterility was recovered. Somatic hybrids were identified morphologically and, for some of them, by cytological observations. They were also characterised by Southern hybridization of nuclear rDNA. Chloroplast and mitochondrial DNA restriction analysis showed that 2 plants out of 14 have B. hirta ctDNA, one the B. napus mtDNA and the other a hybrid. Nine possess B. napus ctDNA with a hybrid mtDNA. For six of them, mtDNA patterns present novel bands, suggesting intergenomic recombination during fusion. These hybrids will be included in the breeding program.
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  • 45
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    Theoretical and applied genetics 75 (1988), S. 833-840 
    ISSN: 1432-2242
    Keywords: Restriction fragment length polymorphism ; Brassica campestris ; Brassica oleracea ; Brassica napus
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The feasibility of creating a restriction fragment length polymorphism (RFLP) linkage map in Brassica species was assessed by screening EcoRI-, HindIII-, or EcoRV-digested total genomic DNA from several accessions of B. campestris, B. oleracea, and B. napus using random genomic DNA clones from three Brassica libraries as hybridization probes. Differences in restriction fragment hybridization patterns occurred at frequencies of 95% for comparisons of accessions among species, 79% for comparisons of accessions among subspecies within species, and 70% for comparisons among accessions within subspecies. In addition, species differences in the level of hybridization were noted for some clones. The high degree of polymorphism found even among closely related Brassica accessions indicates that RFLP analysis will be a very useful tool in genetic, taxonomic, and evolutionary studies of the Brassica genus. Development of RFLP linkage maps is now in progress.
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  • 46
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    Theoretical and applied genetics 76 (1988), S. 165-171 
    ISSN: 1432-2242
    Keywords: Brassica napus ; Somatic hybridization ; Triazine tolerance ; Chloroplast segregation ; RFLP-analysis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Hypocotyl protoplasts of 45 different genotypes of German winter oilseed rape Brassica napus L. (double zero quality: high in yield, seeds low in erucic acid and glucosinolate content) were regenerated to plants. Triazine/triazinone (tri)-tolerant chloroplasts of the Canadian spring oilseed rape variety OAC Triton were introduced into some winter oilseed rapes by means of protoplast fusion. X-ray irradiation was used to limit the transfer of nuclear DNA of Triton protoplasts and to promote the selective transfer of tri-tolerant chloroplasts. Regenerated “cybrid” plants survived a treatment rate of 1000 g/ha metribuzin. The presence and segregation of the tri-tolerant chloroplasts in winter oilseed rape plants, regenerated from fusion products and their progeny, was investigated by restriction fragment length polymorphism (RFLP). Our results indicate that chloroplast segregation was not completed in plants regnerated from fusion products derived from X-irradiated OAC Triton mesophyll protoplasts and German winter oilseed rape hypocotyl protoplasts. In regenerants and their progeny both chloroplast types can still be present. Chloroplasts derived from wintertype protoplasts can outcompete tritolerant chloroplasts during plant development. In some instances, even progeny plants not kept under selective conditions (metribuzin) lost tri-tolerant chloroplasts. A homogenous population of tri-tolerant chloroplasts was necessary to obtain stable tri-tolerant winter oilseed rape plants.
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  • 47
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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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  • 48
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    Molecular genetics and genomics 214 (1988), S. 307-312 
    ISSN: 1617-4623
    Keywords: Brassica napus ; Chloroplast genome ; Anacystis nidulans ; Cyanobacterial transformation ; Plasmid stability
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Brassica napus (cv. “Triton”) chloroplast (cp) DNA BamHI gragments were inserted into a bacteria-cyanobacteria shuttle vector pCB4. The chloroplast genomic library was screened in Escherichia coli and 28 individual clones, which represent 94% of the total chloroplast genome, were isolated. Cyanobacterium Anacystis nidulans R2 was transformed with each member of the clone bank by selection for ampicillin resistance. A study of transformation efficiency showed dramatic variation (up to 200-fold) among recombinant clones. Furthermore, plasmid DNA reisolated from some cyanobacterial transformants exhibited instability. Variations in transformation efficiency and plasmid instability were shown to be DNA sequence specific. B. napus cpDNA clones were thus classified into three types according to their stability in the cyanobacterial host.
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  • 49
    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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  • 50
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    Protoplasma 147 (1988), S. 191-199 
    ISSN: 1615-6102
    Keywords: Brassica napus ; Microspore embryogenesis ; Pollen development
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Populations of highly homogeneous uninucleate and binucleate microspores ofBrassica napus cv. Topas were obtained by bud selection and percoll fractionation. The development of the uninucleate and the binucleate microspores in culture was compared to thosein vivo using the fluorochrome DAPI to stain DNA. The major developmental pathway of the uninucleate microsporesin vitro resulted in embryo formation. The characteristic of this pathway was that the first division produced two diffusely stained nuclei and subsequent divisions gave rise to a multinucleate embryoid. The second pathway which occurred in a small number of the uninucleate microspores led to callus formation. The majority of the binucleate microsporesin vitro followed the developmental pattern of their counterpartsin vivo and were not embryogenic. The embryogenic binucleate microspores produced embryos through the divisions of the vegetative nucleus.
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  • 51
    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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  • 52
    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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  • 53
    ISSN: 1573-5036
    Keywords: Brassica napus ; H ion efflux ; nitrate accumulation ; nitrate reductase ; phosphate nutrition ; xylem exudation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Levels of nitrate reductase activity (N.R.A.) were measured in shoots and roots of P sufficient and P deficient rape plants and changes in N.R.A. examined in relation to the onset of H ion efflux from the roots. Rates of xylem exudation were measured and the sap analysed for nitrate, amino-N and phosphate content. The optimum concentration of phosphate in the leaves for N.R.A. was about 0.7%. Both high and low concentrations of phosphate within the leaves inhibited N.R.A in those leaves. This inhibition of N.R.A led to the accumulation of nitrate in the older parts of the shoots of P sufficient plants. Less accumulation of nitrate occurred in the P deficient plants since nitrate uptake by the plants decreased before any fall in N.R.A. Xylem exudation rates halved within 18 hours of depriving the plants of phosphate, and, since the composition of the sap remained constant, this indicated a reduced flux of nitrate into the xylem. The rate of xylem exudation continued to fall and by the end of the experiment was approximately one tenth of the rate in the P sufficient plants. The onset of H ion efflux from the terminal portions of the root preceded any effect on N.R.A by 2 days.
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  • 54
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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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  • 55
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    Plant and soil 105 (1988), S. 247-256 
    ISSN: 1573-5036
    Keywords: Brassica napus ; cation-anion balance ; H ion efflux ; phosphate nutrition ; root growth
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Changes in pH around the roots of young rape plants were studied using a nutrient film technique which allowed either part or all of the root system to be subjected to specific nutrient treatments. The rapidity and direction of change of pH was assessed by embedding absorbing roots in a thin layer of agar containing bromocresol purple. Measurements were also made with a pH microelectrode placed next to the roots. Phosphate-fed plants were deprived of phosphate when 14 days old. Patterns of pH changes round the deprived roots were the same as with phosphate-fed plants until the plants had been deprived of P for three days, when H ion efflux started in the terminal portions of the roots. The lengths of root producing acid and amounts of H ion both increased as the plants became more P deficient. Both P fed and P deprived roots produced HCO3 ions but the net amount of HCO3 ion produced by the P deficient roots fell as did nitrate uptake rates. Cation-anion balances measured at the end of the experiment showed that uptake of all anions and K decreased in the P deprived plants but uptake of Ca and Mg were little altered. This resulted in a smaller ratio of anions to cations absorbed which was reflected in the reduced HCO3 ion efflux.
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  • 56
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    Euphytica 37 (1988), S. 275-278 
    ISSN: 1573-5060
    Keywords: Brassica napus ; swedes ; vernalisation ; flowering time
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary The effect of pre-vernalisation growth (pre-growth) period on time of flowering was examined in four lines of swedes. No effect was found with the line from cv. Marian. The lines from cvs. Ruta Øtofte and Bangholm Wilby showed increases in time to flowering with increased pre-growth period; maximum delay occurred with 4 wk and 6 wk pre-growth, respectively. Most plants from a line of cv. Criffel showed no effects, but a small proportion did show long delays with pre-growth.
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  • 57
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    Plant and soil 107 (1988), S. 11-18 
    ISSN: 1573-5036
    Keywords: autoradiography ; Brassica napus ; Gossypium hirsutum ; macrostructured soil ; phosphorus depletion ; rhizosphere ; root hair
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract One rape (Brassica napus cv. Wesroona) plant and four cotton (Gossypium hirsutum cv. Sicot 3) plants were grown in plastic cells containing soil labelled with 407 kBq of33P g−1 soil. After 5–8 days of growth, the33P depletion zones of all plants were autoradiographed and33P uptake by plants was measured. The autoradiographs were scanned with a microdensitometer and the optical densities at several places within the33P depletion zones of roots were obtained. The volume of soil explored by root hairs was estimated from measurements of root diameters and lengths of roots and root hairs. About half of the total33P depleted by cotion roots came from outside the root hair cylinder whereas most of33P taken up by rape was from within the root hair cylinder. Plants grown in a macrostructured soil may have roots growing in voids, within aggregates or on the surfaces of aggregates. The results of this study demonstrate that root hairs have a strong influence on the accessibility of phosphorus to roots in such a soil, and thus on the phosphorus nutrition of plants.
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  • 58
    ISSN: 1573-5044
    Keywords: Brassica napus ; protoplasts ; Ficoll ; agarose
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A method is described for regenerating callus from mesophyll protoplasts of a winter variety of Brassica napus. The method combines the use of Ficoll in an initial liquid medium, enhancing early protoplast division and cell colony formation, with a transfer to an agarose system after 10 days culture to give rapid microcalli formation. Further transfers resulted in callus regeneration and the initiation of organogenesis.
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  • 59
    ISSN: 1573-5060
    Keywords: Brassica napus ; B. oleracea ; interspecific hybridization ; first backcross ; triazine resistance
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Atrazine resistant Brassica napus × B. oleracea F1 hybrids were backcrossed to both parental species. The backcrosses to B. napus produced seeds in both directions but results were much better when the F1 hybrid was the pollen parent. Backcrosses to B. oleracea failed completely but BC1s were rescued by embryo culture both from a tetraploid hybrid (2n = 4x = 37; A1C1CC) and sesquidiploid hybrids (2n = 3x = 8; A1C1C). Progeny of crosses between the tetraploid hybrid and B. oleracea had between 25 and 28 chromosomes. That of crosses between the sesquidiploid hybrid and B. oleracea had between 21 and 27. A few plants that had chromosome counts outside the expected range may have originated from either diploid parthenogenesis, unreduced gametes or spontaneous chromosome doubling during in vitro culture. Pollen stainability of the BC1s ranged from 0% to 91.5%. All the BC1s to B. oleracea were resistant to atrazine.
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  • 60
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    Euphytica 38 (1988), S. 277-280 
    ISSN: 1573-5060
    Keywords: Brassica napus ; swedes ; inbreeding ; selection
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary The effect of inbreeding and selection within the old swede cultivar Scotia was examined. A decline in yield was shown by lines produced by selfing from the highest yielding progenies selected after open pollination. It is assumed that these progenies were from heterozygous plants. The yield of two groups of lines did not decline and from these, after a second generation of selfing, eight lines were selected with a mean dry matter yield 10 per cent higher than Scotia.
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  • 61
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    Euphytica 39 (1988), S. 133-136 
    ISSN: 1573-5060
    Keywords: Brassica napus ; swedes ; inbreeding ; selection ; dry matter content
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary A programme of inbreeding and selection was carried out to increase the dry matter content of swedes. A combination of family and individual selection was used, based on deviations from the regression of dry matter content on fresh weight yield. The final selections, from a line of cv. Bangholm Wilby, had a dry matter content of 20 per cent higher than the control cultivars.
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  • 62
    ISSN: 1573-5060
    Keywords: Brassica napus ; Brassica oleracea ; interspecific hybrids ; meiosis ; triazine resistance
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary F1 hybrids of triazine resistant Brassica napus and triazine susceptible B. oleracea were morphologically intermediate to the parent species. Of 49 hybrids examined, 44 had 28 chromosomes, two had 37, one had 38 and two had 56. The 38-chromosome plant was thought to be a matromorph, the others, A1C1C (28), A1C1CC (37) or A1A1C1C1CC (56) type hybrids. Pollen stainability averaged 9.0% in the sesquidiploid, 32.0% in the tetraploids and 89.5% in the hexaploids. All the interspecific hybrids were resistant to 1.0×10-4 mol L-1 atrazine. The sesquidiploid hybrids produced gametes with chromosome numbers ranging from 9 to 17 and the tetraploid hybrid gametes had chromosome numbers from 15 to 22. Most hybrids produced self-seed. The partial fertility of these hybrids may permit their backcrossing to one or both parents.
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  • 63
    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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  • 64
    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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  • 65
    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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  • 66
    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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  • 67
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    Theoretical and applied genetics 73 (1987), S. 465-468 
    ISSN: 1432-2242
    Keywords: Brassica napus ; Resynthesis ; Adaptation to short day ; Earliness
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Natural rapeseed (Brassica napus L.; AACC 2n=38), originated in the temperate climate of the Southwest European Mediterranean region, fails to complete its generative phase in the subtropics and is thus not cultivated in countries like Bangladesh. Adapted agroecotypes are available from the diploid representatives of its genome A (B. campestris/pekinensis, 2n=20) and C (B. oleracea/alboglabra, 2n=18). An artificial resynthesis based on carefully selected progenitor lines was expected to give a photoperiodically better adapted rapeseed. ♀ B. pekinensis x ♂ B. oleracea/alboglabra gave 2 hybrids and 87 matromorphous plants from 1,448 crossed flowers and the reciprocal combination gave no hybrid but 11 matromorphous plants from 2,228 pollinated flowers. The two true hybrids were vegetatively propagated and chromosome doubled. Part of the F2 was grown in Sweden (all plants flowered and the most early ones were selected), part in Bangladesh (13 out of 706 plants flowered). The selected F3 material flowered in Bangladesh and transgressions in earliness could be recorded, some lines were of definite agronomic potential. A correlation in earliness between reaction in Sweden (long day) and Bangladesh (short day) was observed.
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  • 68
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    Theoretical and applied genetics 73 (1987), S. 769-783 
    ISSN: 1432-2242
    Keywords: Microculture ; Electrofusion ; Microinjection ; Karyoplasts ; Cytoplasts ; Protoplasts ; Plant regeneration ; Conditioning ; Brassica napus ; Nicotiana tabacum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Due to the heterogeneity in morphology, physiological and morphogenetical capabilities of higher plant cells in mass culture, the development of methods for individually culturing defined cells seemed to be useful and necessary. Individual cell culture represents a powerful tool for studies on the physiology of different cell types, the analysis of differentiation programs, the genetic manipulation of plant cells and cell-cell interactions. An improved microculture system based on a computer-controlled set-up for the efficient selection, transfer and individual culture of defined higher plant cells until regeneration of whole plants is described. Related experimental approaches for individually manipulating higher plant cells under controlled conditions, such as electrofusion of defined pairs of protoplasts and subprotoplasts, cell reconstruction and intranuclear microinjection of protoplasts and karyoplasts — mainly performed with cells of the crop plant Brassica napus L. — are presented.
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  • 69
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    Molecular genetics and genomics 209 (1987), S. 240-246 
    ISSN: 1617-4623
    Keywords: Brassica napus ; Cybrid formation ; Chloroplasts ; Mitochondria ; MtDNA recombination
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Brassica cybrids were obtained after fusing protoplasts of fertile and cytoplasmic male sterile (CMS) B. napus lines carrying the original b. napus, and the Ogura Raphanus sativus cytoplasms, respectively. Iodoacetate treatment of the fertile line and X-irradiation of the CMS line prevented colony formation from the parental protoplasts. Colony formation, however, was obtained after protoplast fusion. Hybrid cytoplasm formation was studied in 0.5 g to 5.0 g calli grown from a fused protoplast after an estimated 19 to 22 cell divisions. Chloroplasts and mitochondria were identified in the calli by hybridizing appropriate DNA probes to total cellular DNA. Out of the 42 clones studied 37 were confirmed as cybrids. Chloroplast segregation was complete at the time of the study. Chloroplasts in all of the cybrid clones were found to derive from the fertile parent. Mitochondrial DNA (mtDNA) segregation was complete in some but not all of the clones. In the cybrids, mtDNA was different from the parental plants. Physical mapping revealed recombination in a region which is not normally involved in the formation of subgenomic mtDNA circles. The role of treatments used to facilitate the recovery of cybrids, and of organelle compatibility in hybrid cytoplasm formation is discussed.
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  • 70
    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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  • 71
    ISSN: 1617-4623
    Keywords: Agrobacterium ; Ti plasmid ; T-DNA borders ; T-DNA transfer activity ; Plant cell transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We present a detailed analysis of the function of the right and left T-DNA border regions of the nopaline Ti plasmid of Agrobacterium tumefaciens. An avirulent deletion of the right border of the nopaline Ti plasmid (pGV3852) was used as an acceptor for 14 different T-DNA border constructs. The functional activities of these constructs were assayed by their ability to restore virulence, i.e. transformation on inoculated plants. Tumorigenicities were measured in several independent experiments over a 2 year period and the statistical significance of their relative levels was evaluated. The data indicate: (i) the entire sequence of the 25 bp direct repeat of the T-DNA is required to provide an efficient substrate for mediating T-DNA transfer events; deletion derivatives of either the conserved or the vaiable domain of the repeat are defective in T-DNA transfer; (ii) while the 25 bp direct repeat alone can promote the T-DNA transfer, the flanking sequences of the repeats enhance (on the right) or attenuate (on the left) their activity; and (iii) tumorigenicity measurements vary depending on the plant assay system: potato discs are more sensitive than wounded tobacco leaves in detecting differences in T-DNA border activity.
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  • 72
    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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  • 73
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    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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  • 74
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    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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  • 75
    ISSN: 1617-4623
    Keywords: Brassica napus ; Agrobacterium rhizogenes ; Genetic transformation ; Genetic engineering ; Ri T-DNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Genetically transformed repeseed (Brassica napus) roots were obtained by in vitro inoculation of excised stem segments with Agrobacterium rhizogenes. Axenic root organ clones were established and they exhibited a phenotype characteristic of transformed roots: rapid growth, reduced apical dominance and root plagiotropism. Stem regeneration was induced by exposing root fragments to 2,4-dichloroacetic acid (2,4-D) in liquid medium, followed by transfer to solid regeneration medium. The resulting plants exhibited the transformed phenotype observed in other species where similar experiments have been performed. Direct evidence for genetic transformation was obtained from opine assays and molecular hybridization. Sexual transmission of the transformed phenotype was Mendelian, and a probable case of T-DNA insertion into two independent loci within the same plant was detected. The estimated optimal time necessary to obtain transformed oilseed rape plants using this approach is 2 months.
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  • 76
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    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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  • 77
    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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  • 78
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    Molecular genetics and genomics 209 (1987), S. 580-584 
    ISSN: 1617-4623
    Keywords: Rhizobium ; Agrobacterium ; TDNA ; Insertion sequence
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The DNA sequence from a Rhizobium leguminosarum 300 (RL300) plasmid that contains homology to the Tc-DNA of Agrobacterium tumefaciens is described. The RL300 sequence has 78% homology to a 359 bp sequence in the Tc-DNA of pTi15955. The RL300 homology starts approximately 100 bp from the 24 bp border sequence of the TL-DNA and ends approximately 3 bp from an IS66 homolog in the Tc-DNA. An unusual feature of the RL300 homology is the presence of 81 bp direct repeats with Tc-DNA homology, separated by 201 bp. One end of each direct repeat has a 12 bp palindrome. Four cloned sequences of RL300 with homology to the T DNA region were hybridized to plasmid lysates of RL300 derivatives to determine the source of each plasmid. The sequenced homolog, originally on pRH228, was isolated from pRL7JI; the other 3 homologs were isolated from the transmissable plasmids pRL7JI (pRH235) and pRL8JI (pRH235 and pRH236).
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  • 79
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    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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  • 80
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    Euphytica 36 (1987), S. 345-349 
    ISSN: 1573-5060
    Keywords: Brassica napus ; forage rape ; heterosis ; inbreeding depression ; F1 hybrid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Heterosis and inbreeding depression for total fresh and dry weights were studied over two seasons in 15 crosses of six cultivars of forage rape (Brassica napus L.). Some of the hybrids exhibited considerable heterosis over both mid-parent and better parent; followed by depression in the F2. The cross Windal × Canard had 62% and 25% greater dry matter yield over the best cultivar used in this experiment in 1978 and 1979, respectively. Possibilities of the utilization of this hybrid superiority are discussed.
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  • 81
    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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  • 82
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    Plant cell, tissue and organ culture 8 (1987), S. 225-233 
    ISSN: 1573-5044
    Keywords: Brassica napus ; thin cell layer ; protoplasts ; plant regeneration ; rapeseed ; organogenesis ; callus
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Cell layer strips composed of the epidermis and 7–9 layers of subepidermal cells were isolated from the 3–4 terminal internodes of Brassica napus cv Westar plants at the early flowering stage. The strips were precultured for one day in modified liquid MS [11] medium and subsequently incubated for 17–18 h in a 0.4 M mannitol solution containing 1% Macerozyme and 1% Cellulase ‘Onozuka’ R-10. Protoplast yield was 2–2.8×106 per 1.0g of tissue. Protoplasts were cultured at 1×105/ml in three different media: S1 [13], B [12] and L[8]. The first cell divisions occurred after 2–8 days of culture at frequencies of 20–54%. The highest growth rate of colonies was obtained in L medium containing 0.4 M sucrose and 2% Ficoll. After 4 weeks, green calli, 1–2 mm in diameter were transferred onto B5 [2] medium with 3 mgl-1 zeatin, 1% sucrose, 0.1 M mannitol and 0.5% agarose for shoot regeneration. Up to 20% of the calli regenerated shoots which subsequently were rooted and established in soil in the greenhouse.
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  • 83
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    Plant cell, tissue and organ culture 10 (1987), S. 79-90 
    ISSN: 1573-5044
    Keywords: Brassica napus ; B. juncea ; mesophyll protoplasts ; protoplast culture ; regeneration, rapeseed ; mustard
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Various factors affecting the culture of Brassica napus and B. juncea mesophyll protoplasts were examined in order to develop suitable culture media for these species. The basic components (salts and vitamins) of culture media K3 and Kao best supported cell division and colony development in protoplast culture of both species. The addition of casamino acids to Kao's medium resulted in colony browning in B. napus genotypes. B. napus protoplasts grew well with glucose as the osmotic stabilizer, whereas B. juncea protoplasts responded better to sucrose. High NAA and low 2,4-D combinations were effective in stimulating colony growth. Colony development was rapid for a range of genotypes cultured with these recommendations in these media and plant regeneration was obtained from protoplast-derived calli in both species.
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  • 84
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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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  • 85
    ISSN: 1573-5060
    Keywords: Brassica napus ; Brassica oleracea ; interspecific hybridization ; triazine resistance
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Triazine resistant Brassica napus ssp. oleifera and ssp. rapifera were hybridized to cultivars of B. oleracea ssp. italica, ssp. botrytis, ssp. capitata and ssp. fimbriata. The interspecific embryos did not survive in vivo but could be rescued in vitro using a culture medium developed by Monnier (1973). The embryos did not grow directly into normal plants but were successfully regenerated using the protocol developed by Keller (1984). Hybridization efficiency ranged from 0 to 2.64 hybrids per pollination. Interspecific embryo abortion may be related to abnormal endosperm development.
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  • 86
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    Planta 169 (1986), S. 454-461 
    ISSN: 1432-2048
    Keywords: Agrobacterium ; Auxin in tumor tissue ; Crown gall ; Nicotiana (cell transformation) ; Tumor morphology and progression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have isolated two stable variants from a crown-gall teratoma tissue of tobacco (Nicotiana tabacum L.) transformed by Agrobacterium tumefaciens strain A66, a mutant of the virulent A6 strain containing an insertion sequence in the tumor-inducing (Ti) plasmid at the locus coding for auxin biosynthesis. Normally tobacco cells transformed by strain A66 spontaneously form shoots in culture and will not grow on hormone-free medium unless shoots develop. The variant tissue lines, isolated from the teratoma tissue after prolonged culture in the dark, grew as friable and unorganized tissues on hormone-free growth medium. Growth of the variants was more sensitive to auxin feeding than growth of the parental teratoma line, and the auxin dose-response curves of the variant lines were similar to those obtained with A6-transformed tobacco cells. Southern blot analysis of DNA from the parental teratoma line and one of the variants showed no differences in copy number or organization of the oncogenic DNA sequence (T-DNA) transferred from the bacterium, indicating that the variant phenotype did not result from reversion of the A66 mutation. Radio-immunoassay analysis showed similar levels of indole-3-acetic acid (IAA) in the variants and parental teratoma line (3–50 and 38–42 pmol·(gFW)-1, respectively), whereas an A6-transformed cell line contained much higher IAA levels (150–1200 pmol·(g FW)-1). Low levels of the ethylene precursor 1-aminocyclopropane-1-carboxylic acid in the variants and the parental teratoma line (〈5 nmol·(g FW)-1) as compared with that found in the A6-transformed line (〉100 nmol· (g FW)-1) provided additional, indirect evidence for low auxin levels in the variant lines. These results indicate that crown-gall teratoma tissues of tobacco may switch to the unorganized, auxin-sensitive phenotype without an increase in auxin content.
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  • 87
    ISSN: 1432-2048
    Keywords: Agrobacterium ; Brassica (transformed plants) ; Genetic manipulation ; Osmotic pressure ; Potassium ; Solanum (transformed plants)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Growth, water content, osmotic pressure and solute content were examined for normal potato (Solanum tuberosum L. cv. Desiree) and a derivative (line D9X8a), which was genetically transformed with TL-DNA from Agrobacterium rhizogenes. Plants were grown (i) in vitro, (ii) in a growth chamber and (iii) in the field. In vitro, the transformed potato plants produced more biomass than the untransformed plants, partly because they had a higher water content. Potassium concentration and osmotic pressure were lower in cell sap extracted from the transformed potato shoots. In some cases the difference was as much as 50%. These differences were less clear, absent or reversed in plants from a growth chamber or from the field. In the field, however, transformed potato senesced early. It is suggested that a cellular basis for these observations may be changes induced by Ri TL-DNA expression products in plant membrane properties.
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  • 88
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    Theoretical and applied genetics 72 (1986), S. 145-150 
    ISSN: 1432-2242
    Keywords: Brassica napus ; Cytoplasmic male sterility ; Chloroplast DNA ; Mitochondrial DNA ; Restriction patterns
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Restriction patterns of chloroplast (cp) and mitochondrial (mt) DNA in Brassica napus rapeseed reveal the alloplasmic nature of cytoplasmic male sterility in this crop. Both the Shiga and Bronowski systems probably exploit cytoplasmic diversity in B. napus cultivars arising from introgression of cytoplasm from the other rapeseed species, B. campestris. Nuclear genes specific to these systems do not cause sterility in maintainers (Bronowski and Isuzu-natane) because they have a campestris cytoplasm, but give rise to sterility in napus cytoplasms. In the course of hybridization to napus cultivars a line with the triazine resistant cytoplasm (a campestris cytoplasm) has undergone an alteration in the mt genome rendering its restriction pattern more similar than previously to that of napus. The alteration may be an inversion between 7.2 and 3.4 kb in length.
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  • 89
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    Theoretical and applied genetics 72 (1986), S. 373-376 
    ISSN: 1432-2242
    Keywords: Crown gall tumors ; Hairy root disease ; Potato ; Isozyme gene expression ; Agrobacterium ; Solanum tuberosum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Two plant tumors (crown galls and hairy roots) were experimentally provoked on potato cv. ‘Désirée’ by oncogenic strains of Agrobacterium tumefaciens and A. rhizogenes. A marked shift in the expression of some organ-specific genes occurred in crown galls derived from the central zone of tubers: two novel isozyme genes (Est-B and Pox-E) were expressed, two others (Est-C and Pox-F) were suppressed and the remaining ones were maintained in the original state. When the starting tissue was the stem segment, a smaller shift occurred, namely the activation of Adh-A and the suppression of Pox-F. In all cases, the isozyme profiles characterizing all crown galls, whatever their origin, were identical. Under normal aeration conditions, Adh-A was not expressed in either tumoral or non-tumoral roots. However, under the relative anaerobic conditions of in vitro cultures, a difference existed between both types of roots: Adh-A was expressed in normal but not in tumoral roots. This means that hairy roots can tolerate higher levels of anaerobiosis without giving rise to an anaerobic response. For the remaining isozymes, no alteration occurred in either organized (hairy root) or unorganized (crown gall) tumors, as compared to the corresponding non-tumoral tissues (normal root and callus, respectively).
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    Theoretical and applied genetics 71 (1986), S. 716-723 
    ISSN: 1432-2242
    Keywords: Chloroplast DNA map ; Brassica napus ; Triazine resistance ; Canola
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The chloroplast of a triazine resistant weed biotype of Brassica campestris (bird's rape) has been transferred by repeated back-crossing into an agriculturally important strain, ‘Tower’, of Brassica napus to form a triazine resistant cultivar of canola, ‘Triton’, that is low in both erucic acid and thioglucosinolate. In this report, the B. campestris derived chloroplast chromosome of B. napus (cv ‘Triton’) has been cloned into bacterial plasmids and physically mapped for eight restriction enzymes: Apa I, Bam HI, BglI, Hind III, Pst I, Pvu II, Sac I and Xho I. The genes for rRNA, rbcL, cytF, atpA, atpB, atpE, atpH and the triazine resistance gene, psbA were located on the map by heterologous hybridization. The directions of transcription for most of these genes were determined by reverse heterologous hybridization.
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  • 91
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    Theoretical and applied genetics 72 (1986), S. 151-157 
    ISSN: 1432-2242
    Keywords: Brassica napus ; Cytoplasmic male sterility ; Mitochondrial DNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The linear mitochondrial (mt) plasmid in rapeseed occurs in normal, fertile plants, cytoplasmic male sterile (cms) lines (sterile and restored) and maintainers. To determine the role of plasmid genes in cytoplasmic male sterility the three internal Hind III fragments (78%) of the 11.3 kb plasmid were cloned, nick-translated and hybridized to electrophoretic blots of Pst I digests of chloroplast (cp) and mt DNA from normal and cms lines. There is no homology between plasmid DNA and the cp and mt genomes in any line examined nor do plasmid sequences appear to be directly involved in certain alterations of the mt genome. As well, loss of the free plasmid is not associated with integration of the plasmid DNA into the cp or mt genomes. Finally, plasmid copy number appears to be affected by nuclear genotype.
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  • 92
    ISSN: 1617-4623
    Keywords: Agrobacterium ; Ti plasmid ; Ri plasmid ; Opines ; T-DNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Inoculation of carrot discs and Lotus corniculatus plantlets with mixtures of different Agrobacterium rhizogenes or of A. rhizogenes and A. tumefaciens or with Agrobacterium strains harboring both an Ri and a modified Ti plasmid resulted in frequent multiple (pluribacterial) transformation of cells, as revealed by the mixed opine-type of hairy roots arising from them. Multiple transformation may account for the presence of dispersed T-DNA inserts in crown gall and hairy root lines. A plant genetic engineering strategy based on segregation of T-DNA inserts in the progeny of multiple transformants is proposed.
    Type of Medium: Electronic Resource
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  • 93
    Electronic Resource
    Electronic Resource
    Springer
    Molecular genetics and genomics 202 (1986), S. 312-320 
    ISSN: 1617-4623
    Keywords: Agrobacterium ; Plant transformation ; T-DNA borders ; T-DNA transfer
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Crown gall tumors result from transfer and integration of the T-DNA from the Ti plasmid of Agrobacterium tumefaciens into plant nuclear DNA. In the present study, recombinant plasmids containing deletion and rearrangement deriviatives of the T-DNA region of the octopine Ti plasmid pTiA6 were tested in a binary tumorigenesis system (Hoekema et al. 1983) to determine the requirements for T-DNA border regions in tumor formation. Since two defined segments of the T-DNA region of octopine Ti plasmids can be detected in tumor DNA (the left (TL-) and right (TR-) DNA), four border regions exist in this Ti plasmid. Agrobacteria harboring plasmid constructs which contain a T-DNA gene capable of inciting tumors (gene 4, the tmr gene, which is involved in cytokinin biosynthesis) and various T-DNA border regions were tested for ability to cause tumors on Nicotiana glauca and other host plants. Such tmr constructs containing as their only border region the right border of either the TL-DNA or the TR-DNA are fully tumorigenic. Analogous tmr constructs containing only the TL-DNa left border region are not tumorigenic. These results do not depend on the orientation or position of the single border with respect to the tmr gene; furthermore, the TR-DNA right border can confer tumor-forming ability despite the presence of an intervening copy of the TL-DNA left border. These results for relatively small plasmids are contrasted with previously determined requirements for border regions in tumorigenesis by intact Ti plasmids. A model previously proposed by Wang et al. (1984) for the role of border regions in DNA transfer to plant cells is extended in order to explain the tumor-forming ability of plasmid constructs containing a single border region. The results of this study interpreted according to the model suggest that the octopine TL-DNA left border is defective in this DNA-transfer process.
    Type of Medium: Electronic Resource
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  • 94
    ISSN: 1617-4623
    Keywords: Agrobacterium ; Succinamopine ; T-DNA ; AT181
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The T-region of pTiAT181, a SAP(succinamopine)-type Ti-plasmid, is described and is found to be similar to that of the nopaline-type pTiT37. The two major differences are a deletion of 2.9 kb inside the left hand end of pTiAT181, and a right hand region comprised of DNA specific to pTiAT181. This novel DNA was shown by its transfer to plants using a binary vector, to encode an enzyme involved in the synthesis of SAP.
    Type of Medium: Electronic Resource
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  • 95
    ISSN: 1617-4623
    Keywords: Agrobacterium ; Light induction ; Position effect ; Transformed plants ; Gene isolation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The isolation and analysis of several cDNAs and of one genomic clone encoding ST-LS1, a single-copy gene fromSolanum tuberosum with leaf/stem-specific, light-inducible expression, is described. The structure of the gene was determined by sequencing several overlapping partial cDNA clones as well as the genomic clone and by determining the transcription start site by RNA protection experiments. A “tagged” derivative of the gene, obtained by exon modification, was reintroduced into potato and into tobacco shoots usingAgrobacterium/Ti-plasmid vector systems. The modified gene was expressed in both tobacco and potato shoots giving rise to an RNA of approximately 1,200 nucleotides which exceeded the length of the RNA made from the endogenous gene by the expected size of the “tag” (470 nucleotides). The level of expression of the modified gene varied substantially between independent transformants. A high proportion of the transformants (20%–40%) synthesized as much RNA from the added gene as from the resident gene. Expression of the transferred gene was light induced. Qualitatively and quantitatively the expression of the introduced gene was similar in a homologous (potato) and in a heterologous, but related, cellular background (tobacco).
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  • 96
    Electronic Resource
    Electronic Resource
    Springer
    Euphytica 35 (1986), S. 835-842 
    ISSN: 1573-5060
    Keywords: Brassica napus ; Brassica campestris ; rapeseed ; temperature effects ; germination ; germination selection
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Eleven populations of Brassica napus L. and twelve populations of B. campestris L. were subjected to three cycles of selection for fast germination at 2°C and at 25°C. The seeds from the selected populations, and unselected control populations grown in the same environment as the selected populations, were examined for germination behaviour at 2°C and 25°C, and for growth behaviour at 10°C. The populations in both species responded differently in terms of germination behaviour to selection for fast germination. In most of the populations that did respond positively to selection, selection practised at 2°C was superior to selection at 25°C in improving percent germination at 2°C, and was as good as the selection at 25°C in improving germination rate at the higher temperature. Selection for fast germination had no effect on growth characteristics of B. napus and B. campestris populations grown at 10°C. Thus, selection for fast germination at one low temperature may lead to improvement in germination characteristics over a range of temperatures, but will not necessarily lead to improved growth performance of the selected populations.
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  • 97
    ISSN: 1573-5060
    Keywords: Brassica ; Brassica napus ; Brassica juncea ; oilseed crops ; tissue culture ; interspecific hybridization ; genetic variation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Employing in vitro culture of ovaries, ovules and embryos, interspecific hybrids have been obtained amongst two important oilseed crops, Brassica napus x B. juncea and their reciprocal. The test-tube hybrid plants have been transferred to the field, and reared to maturity. The F1 seeds obtained from the hybrid ovaries showed normal germination, and the hybrid plants exhibited a range of variation of characters.
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  • 98
    ISSN: 1573-5036
    Keywords: Brassica napus ; Flowing solution culture ; Growth models ; Hordeum vulgare ; Root growth ; Root hair ; Root length ; Root temperature
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Effects of root temperature on the growth and morphology of roots were measured in oilseed rape (Brassica napus L.) and barley (Hordeum vulgare L.). Plants were grown in flowing solution culture and acclimatized over several weeks to a root temperature of 5°C prior to treatment at a range of root temperatures between 3 and 25°C, with common shoot temperature. Root temperature affected root extension, mean radius, root surface area, numbers and lengths of root hairs. Total root length of rape plants increased with temperature over the range 3–9°C, but was constant at higher temperatures. Root length of barley increased with temperature in the range 3–25°C, by a factor of 27 after 20 days. Root radii had a lognormal distribution and their means decreased with increasing temperature from 0.14 mm at 3°C to 0.08 mm at 25°C. The density of root hairs on the root surface increased by a factor of 4 in rape between 3 and 25°C, but in barley the highest density was at 9°C. The contribution of root hairs to total root surface area was relatively greater in rape than in barley. The changes in root system morphology may be interpreted as adaptive responses to temperature stress on nutrient uptake, providing greater surface area for absorption per unit root weight or length.
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  • 99
    ISSN: 1573-5036
    Keywords: Agrobacterium ; Iron uptáke ; Siderophore
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary An autoradiographic procedure for the investigation of the uptake of minerals by plant roots and its application to the stimulation of uptake of ferric iron by agrobactin is described.
    Type of Medium: Electronic Resource
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  • 100
    ISSN: 1573-5060
    Keywords: Brassica napus ; rape ; heterosis ; sowing density ; physiological correlations ; homozygotes-heterozygotes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Plot yields and both environmental correlations and variances of various morphological and yield characters were compared at different plant densities of lines and F1 hybrids in rapeseed over a two-year period. These variances and correlations were measured on single plant basis and are independent of the genotype: they may partly reflect physiological relations between characters. At intermediate or low density, results indicate no significant heterogeneity for behaviour of both homozygotes and heterozygotes, with some lower correlations at very low densities. At high density hybrid vigour is significantly higher than at low or intermediate density. In heterozygotes, it is associated with reinforced correlations between morphological and yield criteria. The results are discussed with regard to seed yield elaboration, to heterosis and possible homeostasis of hybrids.
    Type of Medium: Electronic Resource
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