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.
出处
《生物技术通讯》
CAS
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)
关键词
橡胶
扩增片段长度多态性
基因组DNA
银染
Hevea brasiliensis
amplified fragment length polymorphism
genome DNA
silver-staining
