发根农杆菌介导的茶树发根高频诱导和遗传转化.pdf

2006 26 11~10 Journal of Tea Science 1, 2 1* 1 1. 310029; 2. 221006 30%OD600 0.5~0.8 60~70 d 10~50 min 100 mmol/L YMB 2 d 500 mg/L MS LG0 PCR Ri T-DNA rolArolB rolC DNA Bt pCAMBIA2301 15834 PCR GUS DNA S571.1; Q523 A 1000-369X200601-001-10 Agrobacterium Rhizogenes-Mediated High Frequency Hairy Root Induction and Genetic Transformation in Tea Plant ZHANG Guang-hui 1, 2, LIANG Yue-rong1*, LU Jian-liang1 1. Department of Tea Science, Zhejiang University, Hangzhou 310029, China; 2. Xuzhou Higher Vocational School of Bioengineering, Xuzhou 221006, China Abstract: An efficient protocol for the establishment of transgenic high hariy root tea plant ( Camellia sinensis) system induction infected by Agrobacterium rhizogenes is reported using two wild type strains of A. rhizogenes , three different co-cultivation media and six different explants. The highest induction frequency was obtained from stem segments of aseptic seedlings infected by A. rhizogenes and co-cultivated on YMB solid media with 100 mmol/L AS for 2 d. The hairy roots grew rapidly on LG0 medium without phytohormone and produced a plenty of branches and root hairs. PCR confirmed that rolA, rolB and rolC have inserted into the genomic DNA of hairy root induced by wild type A. rhizogenes . Hairy roots were also induced from explants of tea plant infected by A. rhizogenes 15834 carrying pCAMBIA2301 binary vector harboring Bt gene, transgenic root were confirmed by PCR and GUS histochemical staining that the Bt exogenous gene was inserted and expressed. Key words: Camellia sinensis, Agrobacterium rhizogenes , hairy root culture, genetic transformation