Abstract:

By Agrobacterium-mediated transformation, a silkworm fibroin light chain gene (FBN), driven by a fiber-specific promoter GAE6-3A, was introduced into the upland cotton G. hirsutum L. cv R15. The plant expression vector used in this study also contains a GUS gene，and a nptⅡ gene driven by 35S promoter, respectively. Kanamycin-resistance analysis, GUS-histochemical staining, and PCR detection were conducted in the 20 T0 generation plants form 11 different transgenic lines. Seventeen plants showed kanamycin resistance, 13 plants were positive in GUS detection, and 14 were positive by PCR analysis for the silkworm fibroin light chain gene FBN. Thirteen positive plants were obtained by the three examination methods among the 20 T₀ generation plants. Southern hybridization of T₁ progenies of lines H18, H21, H32, and H34 were performed, the result showed that among the four T₁ lines, three lines contained two copies and one line contained a single copy of the transgene, no clear hybridization signal was detected for non-transformed control plants, indicating that FBN gene was inserted into the cotton genome by Agrobacterium-medicated transformation. Northern analysis demonstrated that the FBN gene was indeed expressed in the transgenic cotton fiber of above four transgenic lines. Kanamycin-resistance assay and GUS–histochemical staining was used for screening transgenic progenies. Kanamycin assay was performed in the field as a preliminary screening, which may be influenced by climate and other circumstance factors, however, the operation is simple and fast, laying the basis for further selection by GUS histochemical localization. Proceeded the two methods together, the limitation of each measure can be compensated, accordingly the efficiency of homozygous breeding of transgenic progenies could be increased obviously. Till now four T3 positive lines, in addition to several T1 or T2 transgenic plants were obtained, and the foreign gene could be inherited stably form generation to generation. Analysis of fiber quality traits showed that, among the six transgenic lines whose cotton fiber quality was evaluated, the fiber strength of three lines were increased, with the highest in transgenic line H18. These results demonstrate that plant genetic engineering of silkworm fibroin genes has a great potential for improving cotton fiber quality.

Key words: Silkworm fibroin light chain gene, Transgenic cotton, Cotton fiber