太子参ISSR反应体系的优化  

Optimization of the ISSR reaction system for Radix pseudostellarial

作　　者：史辉[1,2] 董晓菲[1] 阮少江[1,2] 叶祖云[1,2] 

机构地区：[1]宁德师范学院生物系,宁德352100 [2]闽东特色生物资源福建省高校工程研究中心,宁德352100

出　　处：《安徽农业大学学报》2016年第5期820-826,共7页Journal of Anhui Agricultural University

基　　金：福建省自然科学基金项目(2016J01169);宁德师范学院"服务海西建设"重点项目(2011H101)共同资助

摘　　要：通过单因子试验和正交设计,对影响太子参ISSR-PCR扩增效果的因素,如模板DNA浓度、Taq酶浓度、dNTPs浓度、引物浓度、退火温度、延伸时间和循环数进行优化。确立了可用于太子参ISSR-PCR分析最适宜的反应体系:20μL反应体系中含80 ng模板DNA,0.5U Taq酶,0.4μmol·L^(-1)引物,0.18 mmol·L^(-1)dNTPs;PCR扩增延伸时间为70 s,循环数为35。稳定性试验表明,该体系能在不同太子参品种中扩增出清晰明亮、稳定性、多态性好的条带。The factors which affected the ISSR-PCR amplification on Radix pseudostellarial such as tem- plate DNA concentration, Taq DNA polymerase, dNTPs, primers, extension time and cycle number were optimized through a combination of single factor and orthogonal design experiments. The results showed that the optimal reaction system for ISSR-PCR of Radix pseudostellarial was as follows： in 20 μL reaction volume, 80 ng template DNA, 0.5 U Taq polymerase, 0.4 μmol·L^-1 primers, 0.18 mmol·L^-1 dNTPs, extended time of 70 s for PCR amplification, and 35 cycles. Stability test results showed that under the optimal system, clear and bright bands with good stability, and polymorphism can be amplified in different varieties of radix pseudostellarial.

关 键 词：太子参 ISSR 反应体系 单因子试验 正交设计 

分 类 号：S567.21[农业科学—中草药栽培]