Yoshiura K I, Kubota T, Soejima H, Tamura T, Izumikawa Y, Niikawa N, Jinno Y
Department of Human Genetics, Nagasaki University School of Medicine, Japan.
Genomics. 1994 Mar 15;20(2):243-8. doi: 10.1006/geno.1994.1160.
To obtain direct evidence for a molecular basis of differentiation between regular Giemsa dark (G)- and light (R)-band regions of human chromosomes, two regions of chromosome 11, q14-q22 (G-band) and q23-q25 (R-band), were microdissected. The DNA fragments were amplified by the linker-primer polymerase chain reaction and cloned into pUC19. Microclones from each library were then compared by colony hybridization with repetitive DNA sequences, by Southern blot hybridization of each microclone to total human genomic DNA and mouse-human hybrid cell DNA containing only human chromosome 11, and by sequencing of unique and Alu-repetitive clones. Among the G-band-derived microclones analyzed, 43.0% were single-copy (unique) sequences and 23.2% contained highly repetitive sequence elements; in the R-band-derived library, 54.2% were unique clones and 20.3% had highly repetitive elements. The G-band library was significantly richer in clones positive for KpnI sequences (4.1% in the G-band library vs 2.8% in the R-band library). No significant difference was found in the proportion of Alu repetitive clones present in the two libraries, but Class IV Alu sequence derivatives were more frequently observed in the R-library than in the G-library. Sequence analysis revealed no significant difference in GC content or in the ratio of CpG sequence to GpC dinucleotide between 30 microclones derived from G-bands and 29 microclones derived from R-bands.(ABSTRACT TRUNCATED AT 250 WORDS)
为了获得人类染色体常规吉姆萨暗带（G带）和亮带（R带）区域分化分子基础的直接证据，对11号染色体的两个区域，即q14-q22（G带）和q23-q25（R带）进行了显微切割。通过连接子引物聚合酶链反应扩增DNA片段，并克隆到pUC19中。然后通过与重复DNA序列的菌落杂交、每个微克隆与总人类基因组DNA以及仅含人类11号染色体的小鼠-人类杂交细胞DNA的Southern印迹杂交，以及对独特克隆和Alu重复克隆的测序，比较了每个文库的微克隆。在所分析的G带衍生微克隆中，43.0%是单拷贝（独特）序列，23.2%含有高度重复序列元件；在R带衍生文库中，54.2%是独特克隆，20.3%有高度重复元件。G带文库中对KpnI序列呈阳性的克隆明显更丰富（G带文库中为4.1%，R带文库中为2.8%）。两个文库中Alu重复克隆的比例没有显著差异，但在R文库中比在G文库中更频繁地观察到IV类Alu序列衍生物。序列分析显示，来自G带的30个微克隆和来自R带的29个微克隆之间，GC含量或CpG序列与GpC二核苷酸的比例没有显著差异。（摘要截短于250字）
