| 大叶栎ISSR扩增体系的优化 |
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| 引用本文： | 梁文汇,谭健晖,黄寿先,梁机,谢素平,刘光金,黄婷,周传明.大叶栎ISSR扩增体系的优化[J].安徽农业科学,2009,37(22):10424-10425. |
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| 作者姓名： | 梁文汇 谭健晖 黄寿先 梁机 谢素平 刘光金 黄婷 周传明 |
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| 作者单位： | 广西大学林学院,国家林业局中南速生材繁育重点实验室,广西南宁,530004;广西林业科学研究院,广西南宁,530001;广西大学林学院,国家林业局中南速生材繁育重点实验室,广西南宁,530004 |
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| 基金项目： | 广西自然科学基金，广西林业科技项目 |
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| 摘 要： | 目的]探讨影响大叶栎ISSR扩增效果的因素，建立大叶栎ISSR扩增的优化体系。方法]以新嫩的大叶栎叶片为试材，采用改良CTAB法提取大叶栎叶片基因组DNA，并将DNA浓度稀释到30ng／μl，采用单因素试验设计测试DNA模板、PCR缓冲液和TaqDNA聚合酶的用量以及底物dNTP终浓度和引物终浓度对ISSR扩增的影响。结果]大叶栎的ISSR扩增的最优反应体系为：总体积20μl中含有DNA模板60ng、PCR缓冲液10×buffer为2．0μl，底物dNTP终浓度为0．25mmol／L，引物终浓度为0．3μmol／L，TaqDNA聚合酶为1U。在该体系下，能够扩增出清晰、多态性高的条带，能够满足大叶栎种群ISSR多样性分析的要求。结论]建立了大叶栎ISSR扩增的优化体系，为研究广西的大叶栎种群遗传多样性提供了技术基础。
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| 关 键 词： | 大叶栎 ISSR 反应体系 优化 |
Optimization of ISSR Amplification System of Castanopsis fissa |
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| LIANG Wen-hui et al.Optimization of ISSR Amplification System of Castanopsis fissa[J].Journal of Anhui Agricultural Sciences,2009,37(22):10424-10425. |
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| Authors: | LIANG Wen-hui |
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| Institution: | LIANG Wen-hui et al(College of Forestry,Guangxi University,Key Laboratory of Fast-growing Tree Breeding in Central South China of State Forestry Administration,Nanning,Guangxi 530004) |
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| Abstract: | Objective] The study aimed to discuss factors that affected ISSR amplification effect of Castanopsis fissa and establish optimized system of ISSR amplification for C. fissa. Method] With tender leaves of C. fissa as tested materials, genomic DNA of C. fissa was extracted with modified CTAB method and the DNA conc. was diluted to 30 ng/μl. Single-factor experiment was designed to test the effect of the dosages of DNA template, PCR buffer and Taq DNA polymerase, as well as the final conc. of substrate dNTP and primers on ISSR amplification. Result] The optimized system of ISSR amplification for C. fissa was as follows: the total 20 μl reaction volume contained DNA template of 60 ng, PCR buffer 10×buffer of 2.0 μl, the final substrate dNTP conc. was 0.25 μmol/L, the final primer conc. was 0.3 μmol/L and Taq polymerase dosage was 1 U. In the system, the clear bands with high polymorphism could be amplified, which could meet the requirements of ISSR diversity analysis for C. fissa. Conclusion] The optimized system of ISSR amplification for C. fissa was established and provided the technical foundation for studying the population genetic diversity of C. fissa population in Guangxi Province. |
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| Keywords: | ISSR Castanopsis fissa ISSR Reaction system Optimization |
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