摘要
以不同白桦个体为材料,通过琼脂糖凝胶电泳法和聚丙烯酰胺凝胶电泳法分析了DNA提取、双酶切、连接、预扩增和选择性扩增过程中的各影响因素,建立了扩增酶切片段长度多态性(Amplified Fragment Length Polymorphism)分析体系,并用E2+M4引物对不同白桦个体进行了扩增,得出优化的关键因素分别为:模板DNA的质量浓度为0.094g/L和0.086g/L,符合AFLP分析中的双酶切要求;酶切反应时间为2h15min;连接最适反应时间为2h;预扩增PCR的退火温度设为60℃。银染结果表明:扩增信号强,无背景干扰,条带清晰。
The influencing factors of several reactions, including DNA extraction, double enzyme digest, adapter ligation, preamplification and selective amplification, were analysed by the methods of agarose gel electrophoresis and polyacrylamide gel electrophoresis using Betula platyphylla as experimental materials. Amplified Fragment Length Polymorphism （AFLP） analysis system established with different birch individuals was performed by primer E2＋ M4. The optimized key factors were obtained as the concentration of template DNA 0. 094 g/L and 0. 086 g/L, 2 hours and 15 minutes for the reaction in the digest system, 2 hours for ligation, and the annealing temperature for pre-amplification 60℃. Clear and strong amplification bands with bright background were obtained after silver staining.
出处
《东北林业大学学报》
CAS
CSCD
北大核心
2007年第5期1-4,9,共5页
Journal of Northeast Forestry University
基金
国家"863"项目(2002BA515B0401)
国家自然科学基金项目(30571513)
东北林业大学校基金资助
关键词
白桦
AFLP
体系优化
Betula platyphflla
Amplified fragment length polymorphism
System optimization
