温郁金SRAP-PCR反应体系建立及条件优化  

Optimization of SRAP-PCR reaction systems for Curcuma wenyujin

作　　者：王晓慧[1,2] 吴亮[2] 姜程曦[2] 

机构地区：[1]吉林省农业科学院农业资源与环境研究所,吉林长春130033 [2]温州医科大学药学院,浙江温州325035

出　　处：《中草药》2016年第14期2515-2518,共4页Chinese Traditional and Herbal Drugs

基　　金：国家"十二五"科技支撑计划(2011BAI04B04);浙江省自然科学基金资助项目(LY15H280014);云南大理药业股份有限公司课题(KJHX1603)

摘　　要：目的研究温郁金Curcuma wenyujin的SRAP-PCR扩增体系最适宜的条件。方法以提取纯化温郁金基因组DNA为模板,利用PCR扩增技术,对SRAP反应中主要影响因素Mg2+浓度、d NTP浓度、模板DNA质量浓度、引物浓度、Taq聚合酶浓度进行优化。结果根据实验确定的最佳体系为25μL SRAP反应体系,其中Mg2+2.0 mmol/L,Taq聚合酶2.0 U,引物浓度1.6μmol/L,模板DNA 0.4 ng/μL,d NTP 2.0 mmol/L,10×PCR缓冲液2.5μL。结论通过扩增条件优化实验,扩增的产物条带清晰明亮、重复性好,确定的反应体系及反应程序适用于温郁金的SRAP分子标记,为今后温郁金的遗传多样性研究奠定了基础。Objective To research on the most appropriate SRAP-PCR amplication system of Curcuma wenyujin. Methods The concentration of Mg2＋, d NTP, template DNA, primer, and Taq polymerase was optimized by amplifying technology and method of PCR based on isolating genomic DNA from C. wenyujin. Results The best amplification system for C. wenyujin was as follows：total reaction volume of 25 μL, including Mg2＋（25 mmol/L） 2.0 μL, Taq polymerase（5 U/μL）0.4 μL, random primer（20 μmol/L）2 μL, template DNA（5 ng/ μ L） 2.0 μ L, d NTP（25 mmol/L） 2.0 μ L, 10 × PCR buffer 2.5 μ L. Conclusion The stripes of amplification products are clearness, high brightness, and good reproducibility. It indicates that the reaction system and reaction procedures which are confirmed in this experiment are applied to the SRAP molecular markers in C. wenyujin, based for the genetic diversity of C. wenyujin.

关 键 词：温郁金 基因组DNA SRAP-PCR反应体系 SRAP分子标记 遗传多样性 

分 类 号：R282.12[医药卫生—中药学]