Expression of a Bacillus thuringiensis toxin (cry1Ab) gene in cabbage (Brassica oleracea L. var. capitata L.) chloroplasts confers high insecticidal efficacy against Plutella xylostella.
Chloroplast genetic engineering is an environmentally friendly approach, where the foreign integrated gene is often expressed at a higher level than nuclear transformation. The cry1Ab gene was successfully transferred into the cabbage chloroplast genome in this study. The aadA and cry1Ab genes were inserted into the pASCC201 vector and driven by the prrn promoter. The cabbage-specific plastid vectors were transferred into the chloroplasts of cabbage via particle gun mediated transformation. Regenerated plantlets were selected by their resistance to spectinomycin and streptomycin. According to antibiotic selection, the regeneration percentage of the two cabbage cultivars was 4-5%. The results of PCR, Southern, Northern hybridization and western analyses indicated that the aadA and cry1Ab genes were not only successfully integrated into the chloroplast genome, but functionally expressed at the mRNA and protein level. Expression of Cry1Ab protein was detected in the range of 4.8-11.1% of total soluble protein in transgenic mature leaves of the two species. Insecticidal effects on Plutella xylostella were also demonstrated in cry1Ab transformed cabbage. The objectives of this study were to establish a gene transformation system for Brassica chloroplasts, and to study the possibility for insect-resistance in dicot vegetables using chloroplast gene transformation.
Liu
Cheng-Wei
CW
Department of Post-Modern Agriculture, Ming Dao University, Chang Hua, 523, Taiwan ROC. chad888@mdu.edu.tw
Lin
Chin-Chung
CC
Yiu
Jinn-Chin
JC
Chen
Jeremy J W
JJ
Tseng
Menq-Jiau
MJ
D000818:Animals; D001413:Bacillus thuringiensis / Q000235:genetics*; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000235:genetics*; D001937:Brassica / Q000382:microbiology*; D002736:Chloroplasts / Q000235:genetics*; D004731:Endotoxins / Q000235:genetics*; D015870:Gene Expression; D005822:Genetic Vectors; D054628:Genome, Chloroplast; D006460:Hemolysin Proteins / Q000235:genetics*; D009036:Moths; D018515:Plant Leaves / Q000235:genetics; D030821:Plants, Genetically Modified / Q000235:genetics*; D018087:Plastids; D014170:Transformation, Genetic; D019076:Transgenes
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D004731:Endotoxins; D006460:Hemolysin Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis