贵州医科大学学报

2013, (03) 231-234+238

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环带库蚊ISSR-PCR反应体系的建立及优化
Establishment and Optimization of an ISSR Reaction System for Culex annulus

修江帆;张春林;李涛;陈汉彬;
XIU Jiangfan,ZHANG Chunlin,LI Tao,CHEN Hanbin(Department of Biology,Guiyang Medical College,Guiyang 550004,Guizhou,China)

摘要(Abstract):

目的:建立及优化适宜环带库蚊(Culex annulus)ISSR-PCR的反应体系,为环带库蚊遗传多样性研究奠定技术基础。方法:选用引物IS01,应用正交试验及单因素试验对的ISSR-PCR反应体系中的BSA、模板DNA、Mg2+、引物、dNTP、Taq DNA聚合酶浓度6个关键因素进行优化筛选。结果:20μL反应体系中最佳因素为BSA 2.0 g/L、DNA模板1.00×10-4mg/L、Mg2+2.0 mmol/L、引物1.0μmol/L、dNTP 0.15 mmol/L、Taq DNA聚合酶0.20 5U/μL。结论:优化后的反应体系适于环带库蚊的ISSR-PCR扩增。
Objective: To establish and optimize an ISSR-PCR amplification system which would provide basis for genetic diversity analysis of Culex annulus.Methods: Primer IS01 were selected to establish the ISSR reaction system.Orthogonal design and single factor method were adopted to screen the optimal conditions for the six critical factors(BSA,DNA template,Mg2+,primer,dNTP,Taq DNA polymerase) of the ISSR-PCR reaction system.Results: The best reaction system was: 20 μL of reaction solution containing BSA 2.0 g/L,DNA template 1.00×10-4 mg/L,Mg2+ 2.0 mmol/L,primer 1.0 μmol/L,dNTP 0.15 mmol/L,and Taq DNA polymerase 0.20 5U/μl.Conclusion: The optimized reaction system is applicable to C.Annulus ISSR-PCR amplification.

关键词(KeyWords): 库蚊属;ISSR-PCR反应体系;遗传多样性
Culex;ISSR-PCR amplification system;genetic diversity

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基金项目(Foundation): 贵阳医学院青年教师科研基金(K2011-39)

作者(Author): 修江帆;张春林;李涛;陈汉彬;
XIU Jiangfan,ZHANG Chunlin,LI Tao,CHEN Hanbin(Department of Biology,Guiyang Medical College,Guiyang 550004,Guizhou,China)

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