薏苡 AFLP体系的建立与优化

摘要

By taking 30 samples of Coix species as testing materials , the genetic diversity and genetic relationship was analyzed using AFLP molecular markers technology .The optimal analysis system of AFLP has been established by studying the key factors of conventional process such as gene extraction , restriction digestion and link up , pre-am-plication, selective amplification, electrophoresis and siliver staining detection .The result showed that the genome DNA was digested more completely by using two-step method compared with one-step method.In each step, the en-zyme reaction time was 4 h added with 15 min for enzyme inactivation .The connection product should be diluted for 10 times and the pre-amplification product should be diluted for 50 times to get a better result .With application of the es-tablished system, 14 selective amplification primers with better polymorphism were selected out from 64 primers.%以30个不同薏苡种质为试验材料，利用AFLP分子标记技术进行遗传多样性及亲缘关系的研究。通过对AFLP常规流程中（基因组提取、酶切、连接、预扩增、选择性扩增、电泳及银染检测）关键因素进行优化，建立适合薏苡的AFLP扩增检测体系。结果表明，采用酶切与连接分开的两步法，酶切完全，每步反应时间4 h并附加15 min的酶失活时间；连接产物稀释10倍；预扩增产物稀释50倍，试验效果最佳。应用优化后体系，从64对引物中筛选出14对多态性较好的薏苡AFLP选择性扩增引物。