短蛸AFLP分子标记分析体系的优化与建立  

The Establishment of AFLP Analysis System for Octopus ocellatus

作　　者：张龙岗[1,2] 杨建敏[1] 刘相全[1] 

机构地区：[1]山东省海洋水产研究所,烟台264006 [2]山东省淡水水产研究所,济南250117

出　　处：《生物技术通报》2010年第5期183-188,共6页Biotechnology Bulletin

基　　金：山东省优秀中青年科学家奖励基金项目(2006BS06010);山东省科技攻关项目(2006GG2205028);国家"863"计划项目(2007AA09Z433)

摘　　要：本研究构建了短蛸扩增片段长度多态性(AFLP)分析体系,对DNA提取、双酶切反应、连接反应、预扩增反应、选择性扩增反应和银染等步骤进行了分析。得到了一种适于短蛸AFLP技术分析的优化体系,该体系中各优化因素为:模板DNA浓度为200 ng/μL;酶切体系中,MseI和EcoR I各加入5 units,缓冲液使用MseI buffer Tango,反应时间为3-4 h;连接最适反应时间为12 h;预扩增产物最适稀释倍数为20倍。该体系的构建为AFLP技术在短蛸分子遗传多样性研究中的应用奠定了基础。AFLP（ amplified fragment length polymorphism）analysis system for Octopus ocellatus was established in this study,with the relative processes being presented, including DNA extraction, double enzymes digestion reaction, adapter ligation reaction,pre-amplification and selective amplification reactions, and argent dyeing. In this paper,some factors affecting the AFLP researching system were studied and an advanced technical system was established. The ascertained factors in the system include concentration of template DNA is 200 ng/μL;5 units of Mse I and EcoR I are added into digest system with Mse I buffer Tango and 3 -4 hours is enough to reaction; the time of ligation should be 12 hours at least;the products of pre-amplification are diluted to 20 times for selective amplification. The establishment of the system might lay a foundation for the application of AFLP techniques to the relative research of Octopus ocellatus.

关 键 词：AFLP 短蛸 反应体系优化 

分 类 号：S917.4[农业科学—水产科学]