蓝莓SRAP—PCR反应体系的建立优化及引物筛选  

Establishment and Optimization of SRAP-PCR System and Primer Screening for Vaccinium ssp

作　　者：尹德洁[1] 苏淑钗[1] 刘肖[1] 侯智霞[1] 陈露[1] 

机构地区：[1]省部共建森林培育与保护重点实验室(北京林业大学),北京100083

出　　处：《东北林业大学学报》2013年第2期35-39,64,共6页Journal of Northeast Forestry University

基　　金：省部级林业公益性行业科研专项(200904014);北京林业大学森林培育科技创新平台开放基金

摘　　要：为进一步利用分子标记技术进行蓝莓品种鉴定及种质资源遗传多样性的分析,拟建立蓝莓新型SRAP标记(相关序列扩增多态性)的优化PCR体系并进行引物组合的筛选。运用改良CTAB法所提取的蓝莓幼嫩叶片的DNA,采用单因子试验,分析了模板DNA浓度、引物浓度、退火温度等影响SRAP-PCR的主要参数,建立了适合蓝莓SRAP-PCR的优化体系,20μL的PCR反应体系:模板DNA浓度120 ng、引物浓度0.40μmol/L、2×TaqPCR Master Mix 10μL,剩余体积用去离子水补足;最佳PCR扩增程序的退火温度为50℃。并从80对SRAP引物组合中筛选出条带清晰且多态性丰富的15对引物组合。The research was conducted for further research of Vaccinium ssp in molecular genetics and marker assistant select breeding by marker technology. An optimized PCR system of SRAP marker （ Sequence-Related Amplified Polymorphism） for Vaccinium ssp and the primer combination of screening were established. The genomic DNA was extracted from the young leaves by modified CTAB method, and three factors affecting SRAP-PCR were analyzed by single-factor test including the concentration of template DNA, primer concentration and annealing temperature. The optimized SRAP-PCR system （ total 20 uL） was established including 0.40 umo]/L primer concentration, 120 ng template DNA, 10 uL 2xTaq PCR Master Mix, and deionized water for making up the remaining volume. Optimal annealing temperature of PCR amplification program was 50℃. 15 primer combinations with distinct and rich polymorphism were screened out from 80 primer combinations of SRAP.

关 键 词：蓝莓 分子标记 SRAP—PCR 体系优化 引物筛选 

分 类 号：S663.9[农业科学—果树学]