Analysis of molecular structure of nucleosomes consisting of repetitive DNA sequences and their function in chromatin

我们研究了在含有（GGA/TCC）n重复序列的DNA片段上重组的单核小体形成DNA-DNA复合物的能力，并通过分离含有GGA重复序列的基因组DNA片段和分析其染色体定位来研究GGA重复序列的功能方面。所得结果如下：1。采用三重亲和力捕获法从小鼠基因组中分离出19个含有（GGA /TCC）n个重复序列的DNA片段。其中19个克隆分别定位在13条染色体的不同位置。含有（GGA/TCC）n重复序列的DNA片段几乎有一半（9/19）富集到异染色质部分，其余部分平均分布在异染色质和常染色质部分。这两个克隆均未富集到常染色质部分。凝胶迁移迁移试验和DNA酶I保护定位结果表明，含有gga重复序列的基因组DNA片段以序列依赖的方式形成DNA- DNA复合物。在含有不同重复数的gga重复序列或在gga重复序列的一个位点或两个位点携带不同侧翼序列的质粒dna片段上重组的单核小体的DNA-DNA复合体的形成分析结果表明，在gga重复序列的两个位点上，gga重复序列的数量和同源侧翼序列都需要通过gga重复序列形成复合体。

We investigated the ability of mononucleosomes reconstituted on DNA fragments containing (GGA/TCC)n repeats for DNA-DNA complex formation and examined the functional aspect of GGA-repeats by isolating genomic DNA fragments containing GGA-repeats and by analyzing their chromosomal localization. The results obtained were as follows.1. We isolated 19 DNA fragments containing (GGA /TCC)n repeats from mouse genome by triplex affinity capture.2. 19 clones of them were mapped to different locations of 13 chromosomes.3. Almost half (9/19) of the isolated DNA fragments containing (GGA/TCC)n repeats were enriched to heterochromatin fraction and the remains were equally distributed to hetero- and eu-chromatin fractions. Neither of the clones examined was enriched to euchromatin fraction.4. The results from gel mobility shift assay and DNase I protection mapping indicated that genomic DNA fragments containing GGA-repeats formed DNA- DNA complexes in a sequence-dependent manner.5. The results from the analysis of DNA-DNA complex formation of mononucleosomes reconstituted on various fragments of plasmid DNAs which contained different repeat-number of GGA-repeats or different flanking sequences carrying at either or both sites of GGA-repeats indicated that their complexes formations though GGA-repeats were required for both GGA-repeat number and homologous flanking sequences at both sites of GGA-repeats.