Isolation of genomic clones homologous to transcribed sequences from human X chromosome

Abstract

Several X chromosome DNA clones homologous to transcribed sequences were isolated from a human X chromosome library. The clones were selected for their ability to hybridize either with³²P -labeled human cDNA in the presence of an excess of unlabeled human repetitive DNA or with mouse fibroblast cDNA. The X chromosome specificity of these sequences was demonstrated by two criteria: A dosage effect was seen when the clones were hybridized to Southern blots of DNA from 1X and 5X cells, and they hybridized to DNA from mouse-human hybrid cells containing only the human X chromosome. The presence of transcribed sequences in these X clones was detected by hybridization with mouse cDNA or with human cDNA in the presence of unlabeled human repetitive sequences, by identifying restriction fragments which hybridize with cDNA but not with human repetitive DNA, and by hybridization with poly A⁺ RNA on Northern blots. These clones were mapped on the human X chromosomes using a panel of mouse-human somatic cell hybrids carrying various translocated human X chromosomes.