Development of insect-resistant transgenic cotton with chimeric TVip3A accumulating in chloroplasts

An optimized vip3A gene, designated as vip3A* was chemically synthesized and a thi1 gene chloroplast transit peptide coding sequence was attached to its 5' end to produce the tvip3A*. vip3A* and tvip3A* genes were transformed into Gossypium hirsutum cv. Zhongmiansuo35 mediated by Agrobacterium tumefaciens. Four independent transgenic T1 lines with single-copy insertions and unchanged phenotypes (CTV1 and CTV2 for tvip3A*, and CV1 and CV2 for vip3A*) were selected by Polymerase chain reaction (PCR), Reverse transcription (RT)-PCR, Southern blotting, enzyme-linked immunosorbent assay (ELISA), and insect bioassay. As expected, the Vip3A* protein of CTV1 and CTV2 were transported to the chloroplasts, where they accumulated. Our results suggest that the two tvip3A* transgenic lines (CTV1 and CTV2) can be used to develop insect-resistant cultivars and could be used as a resource for raising multi-toxins-expressing transgenic cotton.