摘要
对棉花DNA提取程序和SSR反应体系进行了研究。利用正交设计对模板DNA、引物、Taq酶和Mg2+浓度4种成分以3个梯度进行优化和选择,结果表明,最优反应体系为15μL,其中含有:10×Taqbuffer 1.5μL、Mg2+(25 mmol/μL)1.5μLd、NTPs(25 mmol/μL)2.0μL、引物(20 pmol/μL)各2.0μL、Taq酶(2.0U/μL)0.15μL、DNA模板(25 ng)3.0μL、ddH2O补至15μL。采用建立的反应体系,利用8对引物对3个品种进行扩增,6%的变性聚丙烯酰氨凝胶电泳检测结果显示该体系扩增结果清晰稳定。
In this paper the DNA extraction procedure of cotton and the SSR-PCR reaction system were optimized. Using orthogonal design, four factors including template DNA, primer, Taq polymerase,Mg^2＋ at three grads levels were optimized and selected to establish SSR reaction system for cotton molecular breeding. The results indicated that the optimal reaction system were 15 μL. including 10×buffer 1.5μL, Mg^2＋ （25 mmol/μL）l. 5 μL,dNTPs（25 mmol/μL）2.0 μL,Primer （20 pmol/μL）2.0μL for each one, Taq （2.0 U/μL）0.15 /μL,DNA（25 ng）3.0 /μL,ddH2O 2.85 μL. On optimal conditions,9 pairs of primers were used to analyze 3 eultivars of cotton; PCR products were tested by electrophoresis, using 6% denatured polyacrylamide gels. The results showed this optimal system was stable and reliable.
出处
《新疆农业大学学报》
CAS
北大核心
2009年第6期34-37,共4页
Journal of Xinjiang Agricultural University
基金
新疆维吾尔自治区科技厅高技术与发展计划项目(20080101)
新疆维吾尔自治区高校科研计划(XJEDU2008S20)
关键词
棉花
正交试验
PCR
反应体系
cotton
orthogonal design
PCR
reaction system
