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当前位置: 首页 > 文章 > 小麦白粉病菌基因组DNA的微量提取及ISSR-PCR反应体系的优化 植物保护 2010,36 (3) 125-129
Position: Home > Articles > DNA microextraction from wheat powdery mildew and optimization of the reaction system for ISSR-PCR Plant Protection 2010,36 (3) 125-129

小麦白粉病菌基因组DNA的微量提取及ISSR-PCR反应体系的优化

作  者：
朱海荣;段霞瑜;周益林;邱永春
单  位：
中国农业科学院植物保护研究所;沈阳农业大学植物保护学院
关键词：
小麦白粉病菌;DNA微量提取;ISSR-PCR;体系优化
摘  要：
为了寻找适合小麦白粉菌基因组DNA微量提取的方法,本试验利用改进的CTAB法、电钻微量研磨一管法、FastPrep DNA试剂盒法和Chelex-100法分别提取小麦白粉菌基因组DNA。比较得出,在微量提取时,CTAB法的提取率较高;在分生孢子量较多时,FastPrep DNA试剂盒方法提取的DNA质量较高,这两种方法提取的DNA均适用于ISSR-PCR。采用正交设计L16(45)法优化了适合于小麦白粉病菌群体的ISSR体系,确定了25μL时优化的反应体系:1×buffer、模板DNA0.5ng、dNTP0.14mmol/L、TaqDNA聚合酶1U、引物1.4μmol/L、Mg2+1.8mmol/L。利用该体系筛选出了一批多态性较好的ISSR引物,为小麦白粉病菌遗传多样性研究奠定了基础。
译  名：
DNA microextraction from wheat powdery mildew and optimization of the reaction system for ISSR-PCR
作  者：
Zhu Hairong1,2,Duan Xiayu2,Zhou Yilin2,Qiu Yongchun1(1.College of Plant Protection,Shenyang Agricultural University,Shenyang 110161,China;2.Institute of Plant Protection,State Key Laboratory for Biology of Plant Diseases andInsect Pests,Chinese Academy of Agricultural Sciences,Beijing 100193,China)
关键词：
wheat powdery mildew;DNA microextraction;ISSR;optimization of PCR reaction
摘  要：
Four methods were compared to screen a suitable method for DNA microextraction from wheat powdery mildew.A modified CTAB method showed highest extraction rate,and FastPrep DNA kit could also extract high-quality DNA when materials were sufficient.These two methods could be adopted in the analysis of ISSR-PCR.The ISSR-PCR reaction conditions of wheat powdery mildew were optimized by orthogonal design L16 (45).The twenty-five μL amplification reaction system contained 1×PCR buffer,0.5 ng template DNA,0.14 mmol/L dNTP,1 U Taq DNA polymerase,1.4 μmol/L primers and 1.8 mmol/L Mg2+.Some ISSR primers with good polymorphism were screened by using the system,which would provide a basis for further study on genetic diversity of wheat powdery mildew.

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