摘要
[Objective] This study aimed to establish an effective detection technology for rapidly identifying Bt transgenic sugarcane, promptly removing non-Bt transgenic plants and increasing breeding efficiency of Bt transgenic plants, thereby improving breeding efficiency of insect-resistant sugarcane cultivars in China. [ Method] Approximately 1 mg of root tips and other materials were crushed, placed in the bottom of PCR tubes, added successively with solution I and mineral oil, headed at 95 ℃ for 15 rain, added with solution Ⅱ, and finally added with solution III containing GBt primer for PCR. After separated by using electrophoresis on agarese gel, amplification products were observed and photographed under a gel imaging system. [ Result] Various materials treated with different concentrations of solution led to different amplification results. Specifically, the appropriate concentration of NaOH in solution I was 0.1 -0.2 mol/L; the appropriate pH value of solution II was 2. No bands were amplified from mature leaves, tender leaves and old roots of Bt transgenic sugarcane, while root tips successfully amplified 545 bp target bands. The established method was used to identify and analyze 63 hybrid seedlings of YT 91 -976 × SBR216, results showed that a total of 17 samples exhibited 545 bp bands, accounting for approximately 27% of the total number of hybrid seedlings, indicating that these 17 seedlings were Bt transgenic sugarcane plants. [ Conclusion] The rapid detection method established in this study is conducive to identifying Bt transgenic sugarcane and improving breeding efficiency of insect-resistant sugarcane lines, which provides basis for screening of other transgenic materials.
[Objective] This study aimed to establish an effective detection technology for rapidly identifying Bt transgenic sugarcane, promptly removing non-Bt transgenic plants and increasing breeding efficiency of Bt transgenic plants, thereby improving breeding efficiency of insect-resistant sugarcane cultivars in China. [ Method] Approximately 1 mg of root tips and other materials were crushed, placed in the bottom of PCR tubes, added successively with solution I and mineral oil, headed at 95 ℃ for 15 rain, added with solution Ⅱ, and finally added with solution III containing GBt primer for PCR. After separated by using electrophoresis on agarese gel, amplification products were observed and photographed under a gel imaging system. [ Result] Various materials treated with different concentrations of solution led to different amplification results. Specifically, the appropriate concentration of NaOH in solution I was 0.1 -0.2 mol/L; the appropriate pH value of solution II was 2. No bands were amplified from mature leaves, tender leaves and old roots of Bt transgenic sugarcane, while root tips successfully amplified 545 bp target bands. The established method was used to identify and analyze 63 hybrid seedlings of YT 91 -976 × SBR216, results showed that a total of 17 samples exhibited 545 bp bands, accounting for approximately 27% of the total number of hybrid seedlings, indicating that these 17 seedlings were Bt transgenic sugarcane plants. [ Conclusion] The rapid detection method established in this study is conducive to identifying Bt transgenic sugarcane and improving breeding efficiency of insect-resistant sugarcane lines, which provides basis for screening of other transgenic materials.
基金
Supported by Modern Agricultural Industry Technology System (CARS-2004B)
