Taidi-Laskowski B, Tyan D, Honigberg S M, Radding C R, Grumet F C
Department of Pathology, Stanford University School of Medicine, CA 94305.
Nucleic Acids Res. 1988 Aug 25;16(16):8157-69. doi: 10.1093/nar/16.16.8157.
RecA protein-coated probe has been utilized to enrich genomic digests for desired genes in order to facilitate cloning from genomic libraries. Using a previously cloned HLA-B27 gene as the recA-coated enrichment probe, we obtained a mean 108x increase in the ratio of specific to nonspecific plaques in lambda libraries screened for B27 variant alleles of estimated 99% homology to the probe. Class I genes of lesser homology were less enriched: 6.7x for non-B27 genes of estimated greater than 95% homology and 3.7x for other-Class I genes of greater than 80% homology. Loss of genomic DNA during the enrichment procedure can, however, restrict application of this technique whenever starting genomic DNA is very limited. Nevertheless, the impressive reduction in cloning effort and material makes recA enrichment a useful new tool for cloning homologous genes from genomic DNA.
RecA蛋白包被的探针已被用于富集基因组消化产物中的目标基因，以促进从基因组文库中进行克隆。以先前克隆的HLA - B27基因为RecA包被的富集探针，在筛选与该探针具有约99%同源性的B27变异等位基因的λ噬菌体文库中，我们获得了特异性噬菌斑与非特异性噬菌斑比例平均108倍的增加。同源性较低的I类基因富集程度较低：估计同源性大于95%的非B27基因增加了6.7倍，同源性大于80%的其他I类基因增加了3.7倍。然而，当起始基因组DNA非常有限时，富集过程中基因组DNA的损失会限制该技术的应用。尽管如此，克隆工作量和材料的显著减少使得RecA富集成为从基因组DNA中克隆同源基因的一种有用的新工具。
