| 大白菜基因组DNA的提取及AFLP反应体系的建立 |
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| 引用本文： | 孟淑春,张海英,郑晓鹰,刘玉梅,王永健.大白菜基因组DNA的提取及AFLP反应体系的建立[J].分子植物育种,2008,6(2):370-376. |
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| 作者姓名： | 孟淑春 张海英 郑晓鹰 刘玉梅 王永健 |
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| 作者单位： | 1. 国家蔬菜系统工程技术研究中心,北京,100097
2. 中国农业科学院蔬菜花卉研究所,北京,100081 |
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| 基金项目： | 北京市蔬菜种质改良中心建设及产业化开发项目
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北京市自然科学基金 |
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| 摘 要： | 以大白菜嫩叶为材料,利用改进的CTAB法,提取到高质量的大白菜叶片总DNA,通过优化酶切、连接、预扩增、选择性扩增等试验条件,建立了大白菜AFLP银染反应体系,得到了清晰的大白菜AFLP指纹图谱为大白菜品种的分子标记和大白菜品种间亲缘关系等研究奠定了基础.研究结果表明:(1)DNA模板的质量影响酶切以及后续的连接扩增反应,改良的CTAB提取法可用于大白菜AFLP分析,形成清晰的AFLP指纹.(2)大白菜基因组DNA最佳酶切反应体系为:模板DNA总量150ng,反应体积为12.5μL,EcoR I 1.25 U,Mse I 1.25U,5xReaction Buffer,最佳酶切时间为:37℃酶切4h.连接反应于20℃连接2.5h即达最佳效果.(3)6对引物组合(E-AAC/M-CAG、E-AAG/M-CAC、E-ACA/M-CTG、E-ACT/M-CAC、E-ACT/M-CTT、E-ACT/M-CTC)均获得稳定、清晰的扩增图谱,可进行中国大白菜的遗传变异分析.
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| 关 键 词： | 大白菜 DNA提取 AFLP 大白菜 基因组 提取 AFLP 反应体系 Chinese Cabbage Leaves Techniques Establishment 遗传变异分析 中国 扩增图谱 稳定 引物组合 效果 时间 Buffer 体积 最佳 指纹 |
Extraction of DNA and Establishment of AFLP Techniques in Leaves of Chinese Cabbage |
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| Meng Shuchun,Zhang Haiying,Zheng Xiaoying,Liu Yumei,Wang Yongjian.Extraction of DNA and Establishment of AFLP Techniques in Leaves of Chinese Cabbage[J].Molecular Plant Breeding,2008,6(2):370-376. |
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| Authors: | Meng Shuchun Zhang Haiying Zheng Xiaoying Liu Yumei Wang Yongjian |
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| Institution: | Meng Shuchun 1Zhang Haiying 1 Zheng Xiaoying 1 Liu Yumei 2 Wang Yongjian 1 1 National Engineering Research Center for Vegetable,Beijing,100097,2 Institute of Vegetables , Flowers of Chinese Academy of Agriculture Science,100081 |
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| Abstract: | In this study, the improved method of CTAB was used to extract genomic DNA from tender leaves of Chinese cabbage, high quality genomic DNA from leaf of Chinese Cabbage was isolated. DNA restriction reaction, ligase reaction, pre-amplification and selective amplification were optimized for establishment of AFLP reaction system and clear DNA fingerprints. The results indicated: (1) The effect of restriction of DNA and lingation of oligonucleotide adapters and amplification should be influenced by the quality ... |
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