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  • 1
    ISSN: 1573-5028
    Keywords: α-tubulin ; Arabidopsis ; β-glucuronidase ; gene expression ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In Arabidopsis tissues, the pool of tubulin protein is provided by the expression of multiple α-tubulin and β-tubulin genes. Previous evidence suggested that the TUA2 α-tubulin gene was expressed in all organs of mature plants. We now report a more detailed analysis of TUA2 expression during plant development. Chimeric genes containing TUA2 5′-flanking DNA fused to the β-glucuronidase (GUS) coding region were used to create transgenic Arabidopsis plants. Second-generation progeny of regenerated plants were analyzed by histochemical assay to localize GUS expression. GUS activity was seen throughout plant development and in nearly all tissues. The blue product of GUS activity accumulated to the highest levels in tissues with actively dividing and elongating cells. GUS activity was not detected in a few plant tissues, suggesting that, though widely expressed, the TUA2 promoter is not constitutively active.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1573-5028
    Keywords: gene-specific probes ; glutamine synthetase ; transcript accumulation ; Zea mays
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The maize genome has been shown to contain six glutamine synthetase (GS) genes with at least four different expression patterns. Noncoding 3′ gene-specific probes were constructed from all six GS cDNA clones and used to examine transcript levels in selected organs by RNA gel blot hybridization experiments. The transcript of the single putative chloroplastic GS2 gene was found to accumulate primarily in green tissues, whereas the transcripts of the five putative GS1 genes were shown to accumulate preferentially in roots. The specific patterns of transcript accumulation were quite distinct for the five GS1 genes, with the exception of two closely related genes.
    Type of Medium: Electronic Resource
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  • 3
    ISSN: 1573-5028
    Keywords: β-tubulin ; microtubules ; maize ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Four different β-tubulin coding sequences were isolated from a cDNA library prepared from RNA from maize seedling shoots. The four genes (designated tub4, tub6, tub7 and tub8) represented by these cDNA clones together with the tub1 and tub2 genes reported previously encode six β-tubulin isotypes with 90–97.5% amino acid sequence identity. Results from phylogenetic analysis of 17 β-tubulin genes from monocot and dicot plant species indicated that multiple extant lines of β-tubulin genes diverged from a single precursor after the appearance of the two major subfamilies of α-tubulin genes described previously. Hybridization probes from the 3′ non-coding regions of six β-tubulin clones were used to quantify the levels of corresponding tubulin transcripts in different maize tissues including developing anthers and pollen. The results from these dot blot hybridization experiments showed that all of the β-tubulin genes were expressed in most tissues examined, although each gene showed a unique pattern of differential transcript accumulation. The tub1 gene showed a high level of transcript accumulation in meristematic tissues and almost no accumulation in the late stages of anther development and in pollen. In contrast, the level of tub4 transcripts was very low during early stages of male flower development but increased markedly (more than 100 times) during the development of anthers and in pollen. Results from RNAse protection assays showed that this increased hybridization signal resulted from expression of transcripts from one or two genes closely related to tub4. The tub4-related transcripts were not present in shoot tissue. Transcripts from the tub2 gene accumulated to very low levels in all tissues examined, but reached the highest levels in young anthers containing microspore mother cells. RNAse protection assays were used to measure the absolute levels of α- and β-tubulin transcripts in seedling shoot and in pollen. The α-tubulin gene subfamily I genes (tua1, tua2, tua4) contributed the great majority of α-tubulin transcripts in both shoot and pollen. Transcripts from the β-tubulin genes tub4, tub6, tub7, and tub8 were predominant in shoot, but were much less significant than the tub4-related transcripts in pollen.
    Type of Medium: Electronic Resource
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  • 4
    ISSN: 1573-5028
    Keywords: glutamine synthetase genes ; regulation ; nitrate ; ammonium ; Zea mays
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The responses of the five cytosolic-type glutamine synthetase (GS1) genes of maize to treatment of hydroponically grown seedlings with 10 mM KNO3 or 10 mM NH4Cl were analyzed. Non-coding 3′ gene-specific hybridization probes and radioanalytic imaging were used to quantitate individual gene transcript levels in excised roots and shoots before treatment and at selected times after treatment. Genes GS1−1 and GS1−2 exhibited distinct organ-specific responses to treatment with either nitrogen source. The GS1−1 transcript level increased over three-fold in roots, but changed little if any in shoots. In contrast, the GS1−2 transcript level increased over two-fold in shoots, but decreased in roots after treatment. Increased transcript levels were evident at 4 h after treatment with either nitrogen source, with maximum accumulations present at 8 h after treatment with ammonium and at 10–12 h after treatment with nitrate. The GS1−3 gene transcript level showed little or no change after treatment with either nitrogen source. The GS1−4 gene transcript level remained constant in shoots of treated seedlings, whereas in roots, it exhibited relatively minor, but complex responses to these two nitrogen sources. The GS1−5 gene transcript is present in very small amounts in seedlings, making it difficult to analyze its response to metabolites in young plants. These results provide support for the possibility that different cytosolic GS genes of maize play distinct roles in nitrogen metabolism during plant growth and differentiation.
    Type of Medium: Electronic Resource
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  • 5
    ISSN: 1573-5028
    Keywords: gene structure ; introns ; protein homologies ; tubulin ; transcript accumulation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The primary structure of the α1-tubulin gene of Arabidopsis thaliana was determined and the 5′ and 3′ ends of its transcript were identified by S1 nuclease mapping experiments. The information obtained was used to (i) predict the amino acid sequence of the α1-tubulin, (ii) deduce the positions of introns within the α1-tubulin gene, and (iii) construct 3′ noncoding gene-specific hybridization probes with which to study the pattern of α1-tubulin transcript accumulation in different tissues and at different stages of development. The predicted amino acid sequence of the α1-tubulin has 92% identity with the predicted product of the previously characterized A. thaliana α3-tubulin gene. The coding sequence of the α1-tubulin gene is interrupted by four introns located at positions identical to those of the four introns in the α3 gene. RNA blot hybridization studies carried out with an α1-tubulin gene-specific probe showed that the α1 gene transcript accumulates primarily in flowers, with little transcript present in RNA isolated from roots or leaves. In order to investigate the pattern of α-tubulin gene expression in developing flowers, RNA was isolated from flowers at five different stages of development: flower buds, unopened flowers with pollen, open flowers, flowers with elongating carpels, and green seed pods. RNA blot hybridizations performed with 3′ noncoding gene-specific probes showed that the α3 tubulin gene transcript is present in flowers at all stages of development, whereas the α1-tubulin gene transcript could only be detected in RNA from unopened flowers with pollen, open flowers, and flowers with elongating carpels.
    Type of Medium: Electronic Resource
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