Expression of Bacillus thuringiensis (Bt) Crystal Toxin Gene in the Chloroplast of Tobacco
ZHANG Zhong-Lin; REN Yan-Guo; SHEN Yan-Xin; SHAN Song; FAN Guo-Chang'; WU Xiang-Fu'; QMN An-Xian; SHEN Gui-Fang Biotechnology Research Center, Chinese Academy Of Agricultural sciences, Beijing 100081, China Department of Bioscience and Technology, Cheji
The 3.5kb wild-type Bt Cry I A(c) gene and its 3' truncated forms (2.1 kb, l.8kb) were placed under the control of plastid expression signals consisting of the strong light-induced psbA promoter and its 3' untranslated region with the aadA cassette (Prrn, aadA and psbA3' ) as a selectable marker. The resulting vectors PBT3, PBTS and PBT22 also contain flanking tobacco plastid DNA homology regions to direct insertion of the Bt transgene into the tobacco plastid genome between psbA and trnK by homologous recombination. Transformed plastid genomes were selectively amplified by growing the cells on spectinomycin medium. Several independently transformed lines were obtained at last The results of Southern and Western blot demonstrated that these three kinds of Bt genes had been introduced into tobacco plants, and their filial generations are resistant to spectinomycin. Insecticidal activity assay with transgenic tobacco leaves indicate that some plants have strong toxicity to cotton bollworm. This is the first report in China that Bt gene has been introduced and successfully expressed in the chloroplast of higher plants.