柚(Citrus grandis Osbeck)单染色体LA-PCR-AFLP技术体系中外源污染的防控  

Control of Exogenous Contaminations in Pomelo(Citrus grandis Osbeck) Single Chromosome LA-PCR-AFLP

作　　者：王发明[1,2] 王平[1,2] 唐琦[1,2] 柳燕贞[1,2] 陈伟[2] 

机构地区：[1]福建农林大学园艺学院,福建福州350002 [2]福建农林大学生命科学学院,福建福州350002

出　　处：《热带作物学报》2010年第12期2135-2141,共7页Chinese Journal of Tropical Crops

基　　金：国家自然科学基金项目(No.30771488);福建省科技计划项目(No.2007F5018)资助

摘　　要：对柚单染色体LA-PCR-AFLP技术体系中外源DNA污染问题中不同试验用水、加接头连接前后步骤对水污染的敏感度以及LA-PCR形成的非特异性亮带现象进行对比研究。结果表明:8种试验用水中BT水(Bioteke,RNase-free and DNase-free Water)的防控效果最佳;水的质量对LA-PCR过程中接头连接之前的步骤影响最大,对加接头之后的步骤也有一定的影响;非特异性亮带为引物自连形成的多聚体,与外源污染无关。本试验通过对外源污染的来源和关键步骤实施严格防控,经过dot blot验证,使外源污染能够得到有效控制。这为建立高质量的柚单染色体文库奠定了基础。Exogenous DNA contaminations in pomelo（Citrus grandis Osbeck）single chromosome LA-PCR-AFLP were studied by using controlled trials of different types of water,sensitivity test of the water to contaminations before-and-after adding adapter to link,and nonspecific bright bands formed in LA-PCR.The results showed that the BT Water（Bioteke,RNase-free and DNase-free Water）was the most optimum among eight types of test water;Water quality had more important influence before than after adding adapter to link;Nonspecific bright bands were primer polymers which had little to do with exogenous contaminations.Exogenous contaminations could be effectively confined by strictly controlling sources of contamination and strengthening prevention in key steps,and verified by dot blotting.This research has laid foundation for establishing of high-quality pomelo single chromosome libraries.

关 键 词：柚 单染色体 LA—PCR—AFLP 外源污染 DOT BLOT 

分 类 号：S666.3[农业科学—果树学]