Modification of sucrose synthase gene expression in plant tissue and uses therefor
A chimeric gene and plant technology, applied in the field of modification of sucrose synthase gene expression in plant tissue and its application
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Embodiment 1
[0099] Example 1: Inhibition of SuSy Expression in Cotton
[0100] To provide evidence for the role of SuSy in fiber development, the inventors transformed cotton with cosuppressive and antisense SuSy constructs that target the 3' end of SuSy's cDNA and consist of constitutively three Leaf dwarf virus promoter (see Figure 5 for a diagram) drives. The presence of the transgene in 11 transgenic lines was confirmed by Southern analysis (see Figure 6). Two of the antisense lines (294-147, 292-43) and one co-suppression line (295-82) showed a significant reduction in fiber and seed development, while the remaining lines showed varying degrees of inhibition of fiber growth.
[0101] Immunolocalization analysis of 0-d ovule sections (Fig. 2) revealed that SuSy protein in line 294-147 was reduced to less than 20% of wild-type levels compared to wild-type fibroblasts and at 295 -82 down to undetectable. Further, the number and size of initial fibers were reduced by at least 50% in b...
Embodiment 2
[0116] Example 2. Progeny analysis of cotton lines 82, 147 and 43 with suppressed SuSy activity.
[0117] To confirm the correlation between the presence of the SuSy suppressor transgene and the reduction in fibrogenesis in cotton, further analyzes were performed on the T1 generation. Approximately 24 T1 seeds from 6 TO generations were sown. All germinated seedlings were screened by PCR for the presence or absence of this transgene (nptII; S7 promoter and SuSy repressor transgene). Six PCR-positive and one PCR-negative isolates from six strains were analyzed at the molecular, biochemical and cellular levels. Line 82 was vegetatively propagated as a reference point. The fiberless phenotype in different lines of the T0 generation (arrows in FIG. 6A ) was retained in 3 and 2 segregants in lines 147 and 43, respectively (arrows in FIG. 6B ). The remaining individuals showed varying degrees of fiber / seed suppression. Vegetatively-kept line 82 continued to produce fiber-free se...
Embodiment 3
[0121] Example 3. Overexpression of sucrose synthase in cotton plants.
[0122] The coding sequence of a potato sucrose synthase cDNA (Genbank Ogatori No. M18745) was operably linked to a ground clover dwarf virus promoter (S7; WO9606932) and a 3' Transcription termination signal and polyadenylation signal. The chimeric gene is operably linked to a selectable marker gene and introduced into a T-DNA vector. Cotton plants were transformed using the Agrobacterium-mediated transformation technique described above. Transgenic cotton lines were characterized and analyzed for sucrose synthase activity, fiber length, fluff fiber length, cellulose content, and lint dry weight. A certain positive correlation was found between SuSy activity and increases in fiber length, cellulose content and lint dry weight.
[0123] Summary of Examples
[0124] Unicellular cotton fibers arise from the ovule epithelium at anthesis, extend to 2.5-3.0 cm in about 16 days and subsequently synthesize la...
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