| 小麦条锈菌AFLP体系的优化 |
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| 引用本文： | 张妍,康晓慧.小麦条锈菌AFLP体系的优化[J].安徽农业科学,2008,36(6):2227-2228. |
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| 作者姓名： | 张妍 康晓慧 |
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| 作者单位： | 西南科技大学生命科学与工程学院,四川绵阳,621002;西南科技大学生命科学与工程学院,四川绵阳,621002 |
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| 摘 要： | 目的]优化小麦条锈菌AFLP体系。方法]以小麦条锈菌大田混合菌系为材料,采用改良CTAB法提取DNA,探索酶切、PCR过程,优化小麦条锈菌AFLP体系,并从64对引物中筛选出稳定、高效的引物组合。结果]改良CTAB法有较好的破壁效果,所提DNA纯度高,条带清楚,无污染,适合AFLP操作。酶切3.0、3.5 h的效果相同,扩出的片断在100~2 000 bp,可进行预扩。TaqDNA聚合酶为0.5 U时,电泳图中出现了较亮的条带,片断大小在100~2 000 bp。16对引物组合可以扩增出较稳定的条带。结论]优化的小麦条锈菌AFLP体系为:25.0μl酶切体系,37℃酶切3.0 h,10.0μl连接体系在22℃条件下连接过夜,连接产物、预扩产物分别按1∶10、1∶30的比例用ddH2O稀释后用于预扩和选择性扩增。
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| 关 键 词： | 小麦条锈菌 AFLP 反应体系 引物 |
| 文章编号： | 0517-6611(2008)02-02227-02 |
| 修稿时间： | 2007年10月31 |
Optimization of the AFLP System of Wheat Strip Rust |
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| ZHANG Yan.Optimization of the AFLP System of Wheat Strip Rust[J].Journal of Anhui Agricultural Sciences,2008,36(6):2227-2228. |
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| Authors: | ZHANG Yan |
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| Abstract: | Objective] The study aimed to optimize AFLP system of wheat strip rust.Method] With field mixed fungus strains of wheat strip rust as materials,DNA was extracted by improved CTAB method to explore the restriction enzyme digestion and PCR courses and optimize the AFLP system of wheat strip rust.And the stable and high-efficiency primer combinations were screened out from 64 pairs of primers.Result] The improved CTAB method had better effect on breaking cell wall,the extracted DNA had higher purity without pollution and the strip was clear.It was suitable for AFLP operation.The effects of restriction enzyme digestions for 3.0 and 3.5 h were same,the sizes of amplified fragments were in 100～2 000 bp and could be pre-amplified.When Taq DNA polymerase was 0.5 U,the brighter Smear appeared in electrophoretogram with fragment size in 100～2 000 bp.The relatively stable strips could be amplified from the 16 pairs of primer combinations.Conclusion] The optimized AFLP system of wheat strip rust was as follow: digestion with 25.0 μl restriction enzyme system for 3.0 h at 37 ℃,jointing with 10.0 μl joinase system for a night at 22 ℃,using jointing and preamplification products diluted by ddH2O according to the ratio of 1∶10 and 1∶30 resp.in the preamplification and selective amplification. |
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| Keywords: | Wheat strip rust AFLP Reaction system Primer |
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