Essers J, de Stoppelaar J M, Hoebee B
Laboratory of Carcinogenesis and Mutagenesis, National Institute of Public Health and Environmental Protection, Bilthoven, The Netherlands.
Cytogenet Cell Genet. 1995;69(3-4):246-52. doi: 10.1159/000133974.
A DNA segment, containing a so far unknown repetitive DNA sequence has been isolated by reassociation of sheared total rat genomic DNA. FISH with this probe gave a strong hybridization signal on the satellites and in the centromeric region of chromosomes 3 and 12 and on the q-arm of the Y chromosome. A much weaker signal was seen in the centromeric region of chromosomes 11, 19 and X. The repeat unit of this repetitive DNA sequence is a 195-200 bp monomer, which is tandemly repeated. Screening of a rat genomic lambda library resulted in the isolation of variant members of this repeat family. FISH results with these members showed differences in their hybridization pattern especially when posthybridization washings were performed under higher stringency. Under these conditions one phage gave a strong hybridization signal only on the Y chromosome; other phage showed only weak hybridization patterns on different chromosomes. A subclone of the Y-specific phage was sequenced and showed large sequence homology with the 195-200 bp monomer. In Southern blot experiments this Y-specific sequence detects several male specific sequences, although some cross-hybridization with closely related sequences in both male and female DNA can also be observed.
通过剪切的大鼠基因组总DNA重缔合，分离出了一个包含迄今未知的重复DNA序列的DNA片段。用该探针进行荧光原位杂交（FISH），在卫星DNA以及3号和12号染色体的着丝粒区域以及Y染色体的长臂上产生了强烈的杂交信号。在11号、19号染色体和X染色体的着丝粒区域观察到较弱的信号。这个重复DNA序列的重复单元是一个195 - 200碱基对的单体，呈串联重复。对大鼠基因组λ文库进行筛选，分离出了这个重复家族的变异成员。用这些成员进行FISH结果显示，它们的杂交模式存在差异，尤其是在更高严谨度下进行杂交后洗涤时。在这些条件下，一个噬菌体仅在Y染色体上产生强烈的杂交信号；其他噬菌体在不同染色体上仅显示出较弱的杂交模式。对Y特异性噬菌体的一个亚克隆进行了测序，结果显示其与195 - 200碱基对的单体具有高度序列同源性。在Southern印迹实验中，这个Y特异性序列检测到了几个雄性特异性序列，不过在雄性和雌性DNA中也能观察到与密切相关序列的一些交叉杂交。
