棉花BAC文库快速筛选法

A Rapid Method of Screening BAC Library in Cotton

目的:构建棉花细菌人工染色体(Bacterial Artificial Chromosome,BAC)文库的快速筛选法,以期从BAC文库中大量、快速、高效筛选出特定BAC克隆,为从事基因组测序、分离和分析特定基因、构建物理图谱及基因图位克隆等生物学技术研究奠定基础。方法:构建了整板、行、列的三维混合池,以菌液PCR为基础,从BAC文库中筛选出含有特定DNA片段的BAC单克隆。结果:从BAC文库的3 456个克隆中,共筛选出16个阳性单克隆,涉及13条染色体、11个SSR标记。结论:该文构建的棉花BAC文库筛选体系,筛选快速、准确,适合从BAC文库中大量筛选BAC单克隆。结合当前的多种BAC文库筛选方法进行探讨,根据不同的实验目的选择更合适的筛选方法和操作步骤。

Objective:A rapid method of screening BAC library in cotton was constructed to obtain special BAC clones with quality and efficiency,which facilitated biology technology,such as genome sequence,segregation analysis for a gene,construction of physical map,map-based cloning and so on.Method:Three-dimensional pools of plate,row and column were made and bacterial colony PCR was developed to screen BAC clones containing purpose DNA.Result:Sixteen BAC clones were obtained from 3 456 BAC clones,refering to 13 chromsomes and 11 SSR markers.Conclusion:The method was rapid,accurate and appropriate for mass screening.It was also discussed that other methods of screening BAC clones had been used to give a reference for the operators.