Establishing and Optimizing AFLP Amplification Reaction System of Shiraia Bambusicola

文献类型： 外文期刊

第一作者： Yang, Bing

作者： Yang, Bing;Zhou, De-Qun;Yang, Bing;Chen, Yi-Meng;Liu, Yong-Xiang;Liu, Zuo-Yi;Chen, Yi-Meng

作者机构：

关键词： Shiraia Bambusicola;AFLP;System Optimizing;Polymorphic Primer

期刊名称：PROCEEDINGS OF THE 2015 INTERNATIONAL CONFERENCE ON MATERIAL SCIENCE AND APPLICATIONS (ICMSA 2015)

ISSN： 2352-541X

年卷期： 2015 年 3 卷

页码：

收录情况： SCI

摘要： DNA of Sharaia bambuiscola was extracted by the improved method of CTAB. The several key factors affecting the effect of DNA digestion and the PCR selective amplification such as the time of DNA digestion, the times of the per-amplified dilute products, the mount of the selective amplification primer, Taq concentration and dNTPs concentration were optimized and trialed with establishment of an optimized AFLP reaction system of S. bambusicola. The best time of digestion DNA with double endonucleases (EcoR I and Mse I) was 3h. The optimized selection amplification system was 25 mu l reaction mix containing 2.5 mu l the 20 times of the per-amplified dilute products, 2.5 mu L 10x buffer(with Mg2+), 10mmol/L primer each 1.5 mu L, 2.5U.mu L-1 Taq polymerase 0.7 mu L, 10mmol/L dNTP 0.5 mu L, and 15.8 mu L ddH(2)O. Stable and clean DNA finger print can be obtained and 3 pairs of AFLP primers with good genetic diversity were selected according to the optimized reaction system. The results will be an effective protocol for further studying the genetic stucture and differentiation, phylogenetic trees, host specificity and artificial cultivation of Shiraia bambusicola population.

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