Transgenic Brassica napus L. lines carrying a two gene construct demonstrate enhanced resistance against Plutella xylostella and Sclerotinia sclerotiorum
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Plant diseases and insect pests are serious threat to the growth and yield of oilseed rape. In this study, a binary vector carrying sporamin and chitinase PjChi-1 genes in tandem was introduced into Brassica napus cv. ZS 758 via Agrobacterium tumefaciens for dual resistance against disease and insect attack. Thirty-two regenerated plantlets exhibiting hygromycin resistance were selected following Agrobacterium-mediated transformation of 600 leaf petiole explants. Of these, 27 transformants were confirmed to carry the two transgenes as detected by polymerase chain reaction (PCR) with 4.5% transformation efficiency. Eight plantlets were randomly selected for further confirmation by Southern and northern blot hybridization analyses. Four plants carried single copy of the transgenes, while the remaining four plants carried either two or three copies of the transgenes. Moreover, expression of the sporamin transgene was detected by northern blot hybridization in transgenic lines, but not in wild-type plants. These eight T0 plants were grown in vitro, and inoculated with the Lepidoptera larvae of Plutella xylostella and with spores of the fungal pathogen of Sclerotinia sclerotiorum. Transgenic plants exhibited high levels of resistance to P. xylostella and S. sclerotiorum when compared to untransformed wild-type plants. Genetic analysis of T1 progeny confirmed Mendelian segregation of the introduced genes. Therefore, these transgenic lines demonstrate a promising potential for variety development of oilseed rape lines with enhanced resistance against both P. xylostella and S. sclerotiorum.
KeywordsOilseed rape Sporamin Chitinase PjChi-1 Gene expression Insect resistance Antifungal response
This work was sponsored by National Key Science & Technology Supporting Program of China (2010BAD01B01), National Natural Science Foundation of China (30871652, 31000678, 31071698), Industry Technology System of Rapeseed in China (nycytx-005), China National Gene Transformation Program (2008ZX08001-001), and Special Program for Doctoral Discipline of the China Ministry of Education (20090101110102). We thank Drs. Kai-Wun Yeh (Taiwan University), Shusheng Liu and Zhonghua Ma (Zhejiang University) for providing the binary vector, Plutella xylostella larvae and Sclerotinia sclerotiorum pathogen, respectively. The authors are grateful to the editor and referees for their valuable comments to improve our manuscript.
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