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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: To construct a rapid screening method of BAC library to screen a large number of BAC clones from BAC library rapidly, efficiently and to screen specific BAC clones. To perform genomic sequencing, separate and analyze specific genes and construct physical map And gene mapping and other biological cloning lay the foundation for technology research. Methods: A three-dimensional hybrid pool of whole plate, row and column was constructed. Based on bacterial PCR, BAC monoclones containing specific DNA fragments were screened from BAC library. Results: Of the 3 456 clones in the BAC library, 16 positive clones were screened, involving 13 chromosomes and 11 SSR markers. Conclusion: The BAC library screening system constructed in this paper is rapid and accurate. It is suitable for screening large numbers of BAC monoclones from BAC library. Based on the current screening methods of BAC library, we will select more suitable screening methods and procedures according to different experimental purposes.