Labidi B, Broders F, Meyer J L, Hernandez-Verdun D
Université de Paris VI, France.
Biochem Cell Biol. 1990 Jun;68(6):957-64. doi: 10.1139/o90-141.
Investigating genes and their transcription products in nuclear compartments corresponding to one mammalian chromosome, the ribosomal genes 18S-28S and 5S were localized in PtK1 micronucleated cells and rRNA was characterized in sorted micronuclei containing single identified chromosomes. In situ hybridization revealed the presence of 18S-28S rRNA genes in two micronuclei per cell and 5S rRNA genes in four micronuclei per cell. Flow cytometry histograms of isolated micronuclei stained with Hoechst 33342 exhibited five peaks (a-e) in which peaks b and c, respectively, corresponded to chromosomes 4 and X. Restricted genomic DNA from sorted peak c micronuclei showed the presence of 28S gene sequences. Direct sorting of the micronuclei from each peak on nitrocellulose and their hybridization with the 18S-28S rDNA probe revealed that the rRNA genes were exclusively located in micronuclei containing X chromosomes. Northern blotting showed the presence of 18S-28S and 5S rRNAs in peak c micronuclei and their absence from peak b micronuclei. Consequently, these procedures allowed us to show the presence of ribosomal genes and the corresponding rRNA in micronuclei containing single X chromosomes, and the absence of rRNA from micronuclei that do not contain the ribosomal genes. In regards to the transcription of these genes, the micronuclei from peak c can be considered as functional interphase X chromosomes.
在与一条哺乳动物染色体相对应的核区室中研究基因及其转录产物时，核糖体基因18S - 28S和5S在PtK1微核细胞中定位，并且对含有单个已鉴定染色体的分选微核中的rRNA进行了表征。原位杂交显示每个细胞的两个微核中存在18S - 28S rRNA基因，每个细胞的四个微核中存在5S rRNA基因。用Hoechst 33342染色的分离微核的流式细胞术直方图呈现出五个峰（a - eRNA探针杂交显示，rRNA基因仅位于含有X染色体的微核中。Northern印迹显示在峰c微核中存在18S - 28S和5S rRNA，而在峰b微核中不存在。因此，这些方法使我们能够证明在含有单个X染色体的微核中存在核糖体基因和相应的rRNA，而在不含有核糖体基因的微核中不存在rRNA。关于这些基因的转录，峰c的微核可被视为功能性间期X染色体。
