橡胶树AFLP银染体系的建立和优化  

The Establishment and the Optimization of AFLP System for Hevea brasiliensis

作　　者：王惠君[1] 和丽岗[1] 夏志强[1] 王海燕[1] 付瑜华[1] 卢诚[1] 王文泉[1] 

机构地区：[1]中国热带农业科学院热带生物技术研究所热带作物生物技术国家重点实验室,海南海口571101

出　　处：《生物技术通讯》2007年第4期631-634,共4页Letters in Biotechnology

基　　金：国家自然科学基金项目(30571004);国家重点基础研究发展规划项目(2006CB708204)

摘　　要：目的:扩增片段长度多态性(AFLP)为遗传图谱的构建及育种的辅助选择提供了有力的工具。建立一套适合橡胶树的AFLP技术优化体系。方法:以197个GT1×IAN873橡胶树杂交群体为材料,通过对影响AFLP的多种关键因素如模板DNA质量、酶切连接体系、酶切连接反应时间、预扩增体系、选择性体系的分析进行研究。结果与结论:找出一套适于热带植物基因组DNA提取及橡胶树AFLP技术。用改进的CTAB法,经多次抽提纯化,用细玻璃棒挑出DNA得到高质量的模板DNA;酶切连接时DNA模板为250ng,反应体系采用各3U的EcoRⅠ/MseⅡ/T4连接酶,反应时间为9h;预扩增模板为稀释1/2的酶切连接产物,用量为1μL;选择性扩增要用稀释至1/20的预扩增产物。Objective： To establish an optimized amplified fragment length polymorphism（AFLP） system suitable for rub- ber tree. Methods： Some factors affecting the AFLP analysis system for 197 Hevea brasiliensis GT1×IAN873 gemplasms, such as quality of template DNA, the digest-ligage system, the reacting system, the reaction time, preamplification reactions, selective AFLP amplifiction were studied. Results ＆ Conclusion： Find out one set of rubber tree AFLP technology in tropical plants geneome DNA. 250 ng of template DNA in the digest-ligage system, and each 3 U EcoR I/Mse I/T4 ligase at least 9 hours for the reaction time; one microlite digest-ligation products as the template of preamplification 1：2; 40 times of dilution for the products of preamplification with I×TE for selective amplication. The results provided an powerful tool to construct genetic map, the breeding assisted selection.

关 键 词：橡胶 扩增片段长度多态性 基因组DNA 银染 

分 类 号：Q78[生物学—分子生物学]