摘要
以大粒无核葡萄‘皇家秋天’的基因组DNA为模板,对无核葡萄SRAP-PCR反应体系的主要成分及反应退火温度进行优化。获得的最优SRAP-PCR反应程序为94℃预变性5 min,94℃变性1min,38℃复性1min,72℃延伸1min,5个循环;94℃变性1min,56℃复性1 min,72℃延伸1min,35个循环;72℃终延伸10 min。25μL体系中,模板DNA 40ng,Mg2+1.5mmol/L,dNTPs 0.2mmol/L,引物0.2μmol/L,Taq DNA聚合酶0.6U。实验结果表明,优化后的SRAP-PCR反应体系扩增多态性高,带型清晰,稳定性好。
Abstract： In this research, annealing temperatures and several key parameters of the seedless grape SRAP-PCR reaction system were optimized. The result showed that for seedless grape, the most suitable protocol of SRAP-PCR was initially denaturing at 94℃ for 5 min, then 94℃, 1 min, 36℃ 1 min, and 72℃, 1 rain for the first five cycles, and then the annealing temperature was raised to 53℃ for another 35 cycles, at last extending at 72℃ for 10 min. In this SRAP-PCR system, the optimum concentrations of Mg2＋, dNTPs, Taq DNA polymerase, primers and DNA template were 1.5 mmol/L, 0.2 mmol/L, 1.0 U, 0.2 μmol/L, 40 ng/25μL, respectively.
出处
《中国农学通报》
CSCD
北大核心
2011年第16期227-232,共6页
Chinese Agricultural Science Bulletin
基金
国家现代农业产业技术体系建设专项基金(CARS-30-yz-7)
公益性行业(农业)科研专项经费"果树遗传改良与控制技术研究及其应用"(200903044)
陕西省农业科技攻关项目"葡萄新品种引进与标准化生产技术示范"(2009K01-10)
