白芷扩增片段长度多态性反应体系的建立及优化  

Establishment and Optimization of AFLP Analysis System in Angelica dahuruica

作　　者：陈要臻[1] 郭丁丁[2] 马逾英[2] 陈雯[1] 王晓东[1] 唐琳[1] 

机构地区：[1]四川大学生命科学学院,四川成都610064 [2]成都中医药大学中药学院,四川成都610075

出　　处：《时珍国医国药》2008年第12期2844-2846,共3页Lishizhen Medicine and Materia Medica Research

基　　金：国家自然科学基金(No.30572337)

摘　　要：目的建立白芷扩增片段长度多态性(AFLP)反应体系及筛选出扩增条带丰富的引物。方法以12份白芷为试材,对AFLP分析过程中包括提取DNA的方法、DNA的提取质量和浓度、M seⅠ/EcoRⅠ酶切反应时间等进行了研究,从64对引物中筛选适合白芷的引物。结果改良的CTAB方法提取的DNA质量比较好,建立了一种适于白芷AFLP银染技术的优化体系:模板DNA的用量为400 ng,酶切体系中M seⅠ和EcoRⅠ反应时间为4 h,筛选出了7对适合白芷的引物。以引物E+AGC/M+CAA构建的白芷种质资源的AFLP指纹图谱,扩增带多,多态性强。结论建立了适用于白芷的AFLP反应体系,并筛选出了适合白芷的引物,为今后利用AFLP标记技术进行白芷鉴定及种质遗传多样性分析提供了标准化程序。Objective To establish an optimized AFLP （ amplified fragment length polymorphism ） analysis system of Angelica dahuruica. Methods The optimized AFLP analysis system was established with 12 Angelica dahuruica from different habitats after a comparative study of the essential factors which influenced the results of AFLP method. Results The improved CTAB conventional method was better to AFLP than SDS method. The results of the analysis system showed that 1 ） the dosage of template DNA was 400ng; 2） the time of digest system was 4h; 3 ） the products of preamplification should be diluted to 10 times; 4）seven pairs of primer（ E ＋ AGC/M ＋ CAA, E ＋ ACT/M ＋ CAC, E ＋ AGC/M ＋ CAC, E ＋ AAC/M ＋ CAC, E ＋ ACC/M ＋ CAC, E ＋ AAC/M ＋ CAG,E ＋ ACC/M ＋ CTC）were selected from 64 pairs by screening repeatedly. And E ＋ AGC/ M ＋ CAA could amplify more uniform and high sharp bands. Conclusion Our study showed that the AFLP was suitable for genetic diversification, variety identifi- cation,construction of molecular map , identification and classification. These results provide fundamentals for further molecular studies of Angelica dahuruica using AFLP marker.

关 键 词：扩增片段长度多态性反应体系 白芷 

分 类 号：R282.5[医药卫生—中药学]