| Summary: | 碩士 === 中興大學 === 園藝學系所 === 95 === Brassica vegetables play an important role in livelihood of the people and the economic status in Taiwan. In summer, the high temperature and humidity cause very serious and plant diseases in combination with pesis and insects. In order to prevent and control the and insect pest, the farmers spend quite high cost to spray the agricultural chemicals every year in the field.But excessively use of pesticides, not only rsults in resisternce to cheimicals in the insect, also makes a higher cost every year to prevent and conteol the insect. On the other hand, the excessive use of pesticides has caused significant human health problems and environmental damage. Therefore, development of more insectresistant Brassica varieties is considered to be of great economic importance in Taiwan.
Bacillus thuningiensis (Bt) produces a variety of insecticidal crystal proteins (ICPs) upon sporulation which are highly toxic to several insects. Different ICP genes of Bt have been successfully engineered into many crop-plants to obtain resistance against Lepidopteran insects. Chloroplast transformation technologies are a promising tool in biotechnology. On the basis of the increase in activities of multiple Bt proteins providing plants with better insect-defense capability, we propose to transfer cry1Ab, cry1Ac and cry1C genes into the chloroplast of Chinese cabbage. The objectives of this study are to establish the chloroplast gene transfer technology of Chinese cabbage and to study the possibility for improvement of Brassica vegetables with insect resistance and reduce the gene pollution via chloroplast transformation.
The cry1Ab+cry1Ac (pCHL-1Ab-1Ac), cry1Ab+cry1C (pCHL-1Ab-1C), or cry1Ac+cry1C (pCHL-1Ac-1C) genes were transferred into the ''Jade Crown'' cabbage chloroplast via particle gun mediated transformation. Transgenic plants were confirmed by resistance to 10 ppm of spectinomycin. The results of PCR, Southern hybridization, RT-PCR and Western hybridization analysis indicated that the transformed bt genes were integrated into the chloroplast genome and expressed its mRNA and protein. High degree of resistance to the Plutella xylostella were found in the transformed plants containing either one (CCBT-05, expressed cry1Ab) or two bt genes (CCBT-02: expressed cry1Ab and cry1C ; CCBT-05: expressed cry1Ab and cry1Ac).
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