摘要
采用正交和单因素试验设计方法,研究Mg^(2+)、dNTP、引物、TaqDNA聚合酶、模板DNA浓度及退火温度对毛竹ISSR-PCR扩增效果的影响。毛竹ISSR-PCR反应体系为:25μL的体系中含Mg^(2+)1.5 mmol/L,dNTP 0.25 mmol/L,引物1.0μmol/L,TaqDNA聚合酶1.0 U,模板DNA 30 ng,10×Buffer2.5μL。扩增程序为94℃预变性5 min,94℃变性45 s,54℃退火60 s,72℃延伸1.5 min,40个循环;72℃延伸7 min,4℃保存。该体系稳定、可靠,可用于毛竹遗传多样性分析。
The optimal ISSR-PCR system for Phyllostachys heterocycla was established by orthogonal design and single factor test. 25μL reaction system contains 1.5 mmol/L Mg^2＋ , 0.25 mmol/L dNTP, 1.0/.tmol/L primer,1.0 U Taq DNA polymerase,30 ng DNA template, 2.5μL 10 × Buffer. The suitable ISSR-PCR procedure was 1 cycle of pre-denaturing for 5 rain at 94 ℃ ,40 cycles of denaturing for 45 s at 94 ℃, annealing for 60 s at 54 ℃, extending for 1.5 min at 72 ℃, extending for 7 min at 72 ℃, remaining at 4 ℃. The optimized reaction system was suitable for the study of genetic diversity.
出处
《种子》
CSCD
北大核心
2012年第7期28-31,共4页
Seed
基金
广西林业科技项目(编号:桂林科字[2010]第8号)
