Parimoo S, Patanjali S R, Shukla H, Chaplin D D, Weissman S M
Department of Genetics, Yale University School of Medicine, New Haven, CT 06510.
Proc Natl Acad Sci U S A. 1991 Nov 1;88(21):9623-7. doi: 10.1073/pnas.88.21.9623.
Identification of coding segments in large fragments of genomic DNA is a recurrent problem in genome mapping and positional cloning studies. We have developed a rapid and efficient protocol to achieve this goal, based on hybridization of cDNA fragments to immobilized DNA and recovery of the selected cDNAs by the PCR. The procedure permits rapid cloning of cDNA fragments encoded by large genomic DNA fragments, groups of yeast artificial chromosomes, or cosmids and has the potential to directly enrich cDNAs encoded in chromosome segments. By this approach we have been able to identify several non-major histocompatibility complex class I clones from a yeast artificial chromosome that includes the HLA-A locus.
在基因组作图和定位克隆研究中，鉴定基因组DNA大片段中的编码区段是一个反复出现的问题。我们基于cDNA片段与固定化DNA的杂交以及通过聚合酶链反应（PCR）回收所选cDNA，开发了一种快速有效的方法来实现这一目标。该方法能够快速克隆由大基因组DNA片段、酵母人工染色体组或黏粒编码的cDNA片段，并且有可能直接富集染色体区段中编码的cDNA。通过这种方法，我们已经能够从包含HLA - A基因座的酵母人工染色体中鉴定出几个非主要组织相容性复合体I类克隆。
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