Werner D, Neuer-Nitsche B
Institut für Zell- und Tumorbiologie, Deutsches Krebsforschungszentrum, Heidelberg, FRG.
Nucleic Acids Res. 1989 Aug 11;17(15):6005-15. doi: 10.1093/nar/17.15.6005.
A nitrocellulose filter binding assay was applied to isolate and to analyze the fraction of chicken DNA fragments associated with residual nuclear polypeptides resistant to SDS/proteinase K treatment and phenol extraction. It is shown that the DNA-polypeptide complexes retained on nitrocellulose filters are located on a non-random sub-set of DNA sequences. (a) Southern analysis reveals that the fractions of DNA fragments from chicken erythrocytes and from hen oviduct cells associated with the resistant polypeptides have a lower sequence complexity than unfractionated DNA. Moreover, the retained DNA fractions from different cell types of the same species are highly homologous. (b) All DNA fragments of the transcriptionally active and inactive ovalbumin gene map in the DNA fraction passing the filters indicating that the tight DNA-polypeptide complexes are not remnants of transcription complexes. (c) By use of a genomic sub-set library prepared from DNA retained on filters, clones were isolated with sequences mapping specifically in the DNA fraction associated with the tight DNA-polypeptide complexes. The results are consistent with fixed covalent DNA-polypeptide complexes in the chicken genome whose location is essentially identical in different cell types of the same species and apparently determined by DNA signal-sequences.
采用硝酸纤维素滤膜结合分析法来分离和分析与经SDS/蛋白酶K处理及苯酚抽提后仍残留的核多肽相关的鸡DNA片段部分。结果表明，保留在硝酸纤维素滤膜上的DNA-多肽复合物位于DNA序列的一个非随机子集中。(a) Southern分析显示，来自鸡红细胞和母鸡输卵管细胞的与抗性多肽相关的DNA片段部分，其序列复杂性低于未分级的DNA。此外，来自同一物种不同细胞类型的保留DNA部分高度同源。(b) 转录活性和非活性卵清蛋白基因的所有DNA片段都位于通过滤膜的DNA部分中，这表明紧密的DNA-多肽复合物不是转录复合物的残余物。(c) 通过使用从保留在滤膜上的DNA制备的基因组子集文库，分离出了一些克隆，其序列特异性地定位在与紧密DNA-多肽复合物相关的DNA部分中。这些结果与鸡基因组中固定的共价DNA-多肽复合物一致，其位置在同一物种的不同细胞类型中基本相同，并且显然由DNA信号序列决定。
