| 百脉根ISSR-PCR反应体系的建立与优化 |
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| 引用本文： | 富新年,师尚礼,王赞,李俊,闫洪唯.百脉根ISSR-PCR反应体系的建立与优化[J].草原与草坪,2012,32(3):49-52,57. |
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| 作者姓名： | 富新年 师尚礼 王赞 李俊 闫洪唯 |
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| 作者单位： | 1. 甘肃农业大学草业学院／草业生态系统教育部重点实验室／甘肃省草业工程实验室／中-美草地畜牧业可持续发展研究中心,甘肃兰州,730070
2. 中国农业科学院北京畜牧兽医研究所,北京,100193 |
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| 基金项目： | 国家现代牧草产业技术体系，牧草种质资源保护与利用，十二五科技支撑 |
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| 摘 要： | 运用L16(45)正交设计对百脉根ISSR反应的5个因素,即DNA模板浓度、dNTPs浓度、引物浓度、Mg2+浓度、TaqDNA聚合酶浓度在4个水平上进行优化实验,通过不同反应体系扩增效果比较,最终确定在20uL体系中各反应物的最适含量为:40ng模板DNA、2μL 10×PCR Buffer、0.15mmol/LdNTPs、0.75μmol/L ISSR引物、1.5mmol/L MgCl2、1UTaqDNA聚合酶。该体系的建立为今后利用ISSR技术进行百脉根属种质资源的遗传多样性分析奠定了技术基础。
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| 关 键 词： | 百脉根 ISSR 反应体系 优化 |
Establishment and optimization of ISSR reaction system for Lotus corniculatus |
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| FU Xin-nian,SHI Shang-li,WANG Zan,LI Jun,Yan Hong-wei.Establishment and optimization of ISSR reaction system for Lotus corniculatus[J].Grassland and Turf,2012,32(3):49-52,57. |
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| Authors: | FU Xin-nian SHI Shang-li WANG Zan LI Jun Yan Hong-wei |
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| Institution: | 1.College of Pratacultural Science,Gansu Agricultural University/Key Laboratory of Grassland Ecosystem,Ministry of Education/Pratacultural Engineering Laboratory of Gansu Province/Sino-U.S. Centers for Grazingland Ecosystem Sustainability Lanzhou 730070,China; 2.Institute of Animal Science,Chinese Academy of Agricultural Sciences,Beijing 100193,China) |
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| Abstract: | The orthogonal design of five factors,concentrations of DNA template,dNTPs,primer,Mg2+and Taq DNA polymerase,in four levels was used to optimize ISSR-PCR amplification system for Lotus corniculatus.The results showed that the optimum concentration of seven reactants in 20 μL reaction mixture were as follows:40 ng genomic DNA,2 μL 10 × PCR buffer,0.15 mmol ·L-1 dNTPs,0.75 μmol ·L-1 ISSR primer,1.5 mmol ·L-1 MgCl2,1 U Taq DNA polymerase.The reaction system would provide the basis for the diversity analysis of Lotus corniculatus with ISSR markers. |
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| Keywords: | Lotus corniculatus ISSR reaction system optimization |
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