摘要
以大豆为试材,研究了大豆SSR检测体系中PCR扩增的各主要成分对检测结果的影响,确立了适宜的退火温度和反应体积,改进了PCR扩增程序和银染方法,进一步优化了适用于大豆的SSR检测体系。优化后的SSR反应体系为:10×Buffer 1.00μL、2.00 ng/μL模板DNA、0.15μmol/L SSR引物、150.00μmol/L dNTPs、0.50 U Taq酶、2.00mmol/L Mg2+,加ddH2O至终体积10.00μL。优化的PCR扩增程序为:94℃预变性3 min;94℃变性25s,47℃退火25s,72℃延伸30s,共35个循环;72℃延伸5min,4℃保存。优化后的检测体系更为经济有效。
Effects of major components of PCR amplification on soybean SSR detection results were studied. A SSR detection system suitable for soybean was established with improved PCR reaction system, PCR program and silver staining method. The optimized PCR amplification system was 10× Buffer 1. 00μL, 2. 00 ng/μL template DNA, 0. 15 μmol/L SSR primer, 150. 00μmol/L dNTPs, 0.50 U Taq enzyme, 2.00 mmoL/L Mg2＋ , adding ddH2O to terminal volume 10.00μL. The optimized PCR cycles started at 94℃ （3min） ,followed by 35 cycles of 25s at 94℃, 25s at 47℃ and 30s at 72℃, and then extended 5min at 72℃. The optimized soybean SSR detec- tion system was proved more economic and useful.
出处
《大豆科学》
CAS
CSCD
北大核心
2007年第1期36-40,共5页
Soybean Science
基金
辽宁省教育厅科技项目(05L3781)
辽宁省科学技术计划项目(2006201008)的资助
关键词
大豆
SSR
检测体系
优化
Soybean
SSR
Detection system
Optimization
