A novel method for the genetic transformation of cotton pollen by means of vacuum infiltration and Agrobacterium-mediated transformation is reported. The acsA and genes, which are involved in cellulose synthesis in Acetobacter xylinum, were transferred into pollen grains of brown cotton with the aim of improving its fiber quality by incorporating useful prokaryotic features into the colored cotton plants. Transformation was carried out in cotton pollen-germinating medium, and transformation was mediated by vector pCAMBIA1301, which contains a reporter gene β-glucuronidase (GUS), a selectable marker gene, hpt, for hygromycin resistance and the genes of interest, acsA and acsB. The integration and expression of acsA, acsB and GUS in the genome of transgenic plants were analyzed with Southern blot hybridization, PCR, histochemical GUS assay and Northern blot hybridization. We found that following pollination on the cotton stigma transformed pollen retained its capability of double-fertilization and that normal cotton seeds were produced in the cotton ovary. Of 1,039 seeds from 312 bolls pollinated with transformed pollen grains, 17 were able to germinate and grow into seedlings for more than 3 weeks in a nutrient medium containing 50mg/l hygromycin; eight of these were transgenic plants integrated with acsA and acsB, yielding a 0.77% transformation rate. Fiber strength and length from the most positive transformants was 15% greater than those of the control (non-transformed), a significant difference, as was cellulose content between the transformed and control plants. Our study suggests that transformation through vacuum infiltration and Agrobacterium mediated transformation can be an efficient way to introduce foreign genes into the cotton pollen grain and that cotton fiber quality can be improved with the incorporation of the prokaryotic genes acsA and acsB.

A glutathione S-transferase (GST) gene has been introduced into restorers of cytoplasmic male sterility in upland cotton (Gossypium hirsutum L.) using Agrobacterium-mediated transformation. A transgenic restorer, signed as 'Zheda strong restorer', which has strong restorability to male sterility, was selected from progeny plants of transformants. When compared with an American restorer 'DES-HAF277', the fertility restorability of 'Zheda strong restorer' to male sterility was been enhanced by 25.8% in the percentage of viable pollens of hybrid (sterile line

A mRNA preferentially expressed in cotton fiber was cloned from fiber total RNA of normal upland cotton TM-1 (wild-type) by using RT-PCR and corresponding cDNA (signed as TM-E6) was sequenced. TM-E6 gene had no intron and contained an open reading frame 771 bp long, and might encode a peptide of 246 amino acids. Other four genes, Fl-E6, Li-E6, N-E6 and Bl-E6, which were homologous to TM-E6 gene, also were isolated from four fiber mutants of Fiberless Xu-Zhou 142, Ligon lintless, Naked seed and Brown lint, respectively. Sequence analysis of each of these mutant genes revealed many variations in structure and nucleotide composition of gene when compared with sequence of TM-E6 gene. (1). There was a changeable repetitive segment in which GGCTCA (Gly-Ser) was repeated from three to five times between the 82nd and the 93rd codon in different mutant genes. Since the change of Gly-Ser repetitive segment occurred not only in the mutants but also in the wild-type cotton, the repeat frequency might be not associated with the mutation of fiber characteristics. (2). Among these four mutant genes, the percentage of changed codons was 7.05% in Fl-E6, 4.98% in Li-E6, and 4.15% in N-E6 and Bl-E6. It seems that the percentage of changed codons in E6 sequence was positively correlated to the degree of fiber morphological variation. (3). E6 polypeptides of Two long-fiberless mutants (Fiberless Xu-Zhou 142 and ligon lintless) contained high similar (99.4%) variation in the region of ~174 amino acids from N-terminus, and those of shor-fiberlees mutants (Fiberless Xu-Zhou and naked seed) revealed identical variation in the region from 116th to 220th amino acid. It also seems that there was a parallel relation between E6 protein variation and fiber phenotype mutation. (4). Li-E6 and Bl-E6 gene also expressed at low level in seed coat besides at high level in fiber.

In the present study, three types of color fiber cottons i.e. white, brown and green were compared for their fiber quality and yield. The comparison of fiber quality suggested that color fiber cotton was inferior as compared to white fiber cotton. To know the effect of cellulose, mineral elements (N, P and K) and pH of fiber cells on quality of fiber, these components were studied at different fiber cell development stages in all the three fiber cotton types. The cellulose content is closely associated with quality of fiber. The higher fiber quality of white fiber cotton might be due to the high cellulose content in it as compared to colored fiber cotton. A rapid and slow decrease in pH in white and colored cottons, respectively, might bring some effects in fiber elongation. Among the mineral contents, Potash (K) element positively correlated with the fiber quality traits. The pigment development in brown and green fiber cottons was not similar. In green fiber cotton it take more time to become deep as compared to brown fiber cotton. Ultimately, the possible strategy for improvement in quality of color fiber cotton was discussed. The results of heterosis in colored fiber cotton suggested that heterosis approach could improve yield and quality of colored fiber cotton. In the present study, the hybrid between ZJU12A

A fiberless seed mutant (fl) was identified in a commercial cotton (Gossypium hirsutum L.) variety Xu-Zhou 142 (FL). This phenotype is associated with lack of fiber cell initiation in the outer integument of the ovule, as was characterized by analysis of genes related to fiber differentiation and development. Two genes, fl-E6 and FL-E6, were cloned from fl-integument cells and FL-fiber or integument cells, respectively. Compared with FL-E6, fl-E6 showed a dramatic change in nucleotide sequence: (1). FL-E6 contained a tandem repetitive sequence in which GGCTCA (Gly-Ser) is repeated five times between the 82nd and the 93rd codon from the first ATG codon, while in fl-E6 the same sequence is repeated four times;(2). The fl-E6 gene encodes a polypeptide of 241 amino acids but lacks two codons between the 90th and 93rd codon and three between the 171st and 174th relative to FL-E6;(3). There are also 12 nucleotide substitutions which would result in 7 amino acid differences between fl-E6 and FL-E6. Analysis of RT-PCR and Northern Blot showed that expression of the fl-E6 gene is suppressed in the fl-integument cells, but highly expressed in FL-fiber cells. The difference between fl-E6 and FL-E6 may be associated with lower expression of fl-E6 in the fl-integument cells. Searches of protein databases with the FL-E6 gene sequence showed similarity to the protein backbones of two arabinogalactan-proteins (AGPs), one from the filtrate of suspension-cultured cells of Pyrus communis (AGPPc2) and the other from Nicotiana alata (AGPNa2). Although the function of the FL-E6 protein in differentiation and development of cotton fiber cells is not known, the data indicate that the mutation of fl-E6 gene from FL-E6 gene may inhibit the fiber cell initiation from epidermal cells of the outer integument of the ovule.