摘要
以秦岭野生大叶铁线莲为试验材料,在单因素试验的基础上进行L9(34)正交试验,从而优化大叶铁线莲ISSR-PCR反应体系。结果显示,在25μL反应体系中,最佳体系为1.0μL模板DNA、2.0μL Mg^2+、2.0μL dNTPs、1.0μL引物以及0.2μL Taq DNA聚合酶和18.8μL ddH2O。最佳扩增程序为:94℃预变性5 min;94℃变性50 s,47℃退火1 min,72℃延伸1 min,35个循环;72℃延伸10 min,最后4℃保存。
In this experiment,the wild Clematis heracleifolia DC.was used as the experimental material in the Qinling mountains.Based on the single factor experiment,the L9(34)orthogonal test was carried out to optimize the ISSR-PCR reaction system of Clematis heracleifolia DC.The results showed that in the 25μL reaction system,the optimal system was 1.0μL of template DNA,2.0μL of Mg^2+,2.0μL of dNTPs,1.0μL of primer,0.2μL of Taq DNA polymerase,and 18.8μL of ddH2O.The optimal amplification procedure was,pre-denaturation 94℃for 5 min,denaturation 94℃for 50 s,annealing 47℃for 1 min,extension at 72℃for 1 min,35 cycles,extension at 72℃for 10 min,and storage at 4℃.
作者
任夏萌
李欣怡
丁群英
REN Xiameng;LI Xinyi;DING Qunying(College of Biological and Environmental Engineering,Xi'an University,Xi'an 710000,China)
出处
《山西农业科学》
2019年第12期2083-2087,共5页
Journal of Shanxi Agricultural Sciences
基金
国家级大学生创新创业训练计划项目(201811080015)
西安市科技计划项目(2019KJWL16)
