摘要
利用正交试验设计的方法,从Mg2+浓度、dNTPs浓度、引物浓度和Taq酶浓度4因素对菊芋ISSR-PCR反应体系进行优化分析,并在此基础上对模板DNA浓度及退火温度进行梯度检测。结果表明:菊芋20μl最佳反应体系包括10×PCR buffer,200μmol/L dNTP,0.5μmol/L引物,1.5 mmol/L Mg2+,1.0 U Taq DNA聚合酶和50 ng模板DNA。这一优化系统的建立,将为菊芋种质资源鉴定及遗传多样性的研究奠定基础。
In order to optimizing ISSR-PCR reaction system in Helianthus tuberosus L.,the concentrations of Mg2＋,dNTPs,primer DNA,and Taq DNA polymerase were optimized at three levels by orthogonal design.Then,based on the optimal ISSR-PCR amplification the concentration of DNA template and annealing temperature were selected.As a result,the optimal PCR（20 μl）mixture contained 10×PCR Buffer,200 μmol/L dNTP,0.5 μmol/L primer,1.5 mmol/L MgCl2,1.0 U Taq polymerase and 50 ng DNA template.This optimized system would play an important role in further research on germplasm identification and the genetic diversity of H. tuberosus by using ISSR molecular marker techniques.
出处
《西南农业学报》
CSCD
北大核心
2012年第1期243-246,共4页
Southwest China Journal of Agricultural Sciences
基金
国家大宗蔬菜产业技术体系西宁综合试验站(CARS-25-G-49)
青海省蔬菜遗传与生理重点实验室创新基金(Sc-zdsys-2011-02)
关键词
菊芋
ISSR-PCR
优化
Helianthus tuberosus L.
ISSR-PCR
Optimization
