柚(Citrus grandis Osbeck)单染色体LA-PCR-AFLP分子标记体系的建立  

Establishment of Single Chromosome AFLP System in Pomelo (Citrus grandis Osbeck)

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10期1778-1784,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：通过微分离和微克隆技术,对柚单染色体LA-PCR-AFLP技术体系中反应程序、模板浓度等关键参数进行了优化和引物筛选。结果表明:退火温度较高的程序TD3[退火梯度:72℃→65℃30 s(-0.5℃/cyc)]扩增效果较好;稀释100倍(10 ng/μL)为最适模板量;10个引物对组合扩增多态性较好。对22条单染色体的AFLP多态性分析和dot-blot杂交验证,表明较好地构建了柚单染色体LA-PCR-AFLP体系。这为柚单染色体的识别和文库构建提供了试验基础。The key parameters of LA-PCR-AFLP technical system were optimized for available PCR protocol,optimum template concentration and better primer pairs on the base of chromosome microdissection and microcloning technique.Results showed that program TD3 with higher annealing temperature [72 ℃ 65 ℃ 30 s（-0.5 ℃/cyc）] had better amplification effect;the optimal template amount was diluted with 100 times（10 ng） and 10 primer pairs had better amplified polymorphism.AFLP polymorphism analysis of 22 single chromosomes and dot-blot hybridization proved that the single chromosome LA-PCR-ALP technical system in Pomelo was preferably established,which offered an experimental basis for the identification of homologous chromosomes and the construction of library

关 键 词：柚 单染色体 AFLP体系建立 dot—blot 

分 类 号：S666.3[农业科学—果树学]