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克隆仓鼠端粒处的一种新型小卫星。

A novel minisatellite at a cloned hamster telomere.

作者信息

Shampay J, Schmitt M, Bassham S

机构信息

Department of Biology, Reed College, 3203 SE Woodstock Blvd., Portland, OR 97202-8199, USA.

出版信息

Chromosoma. 1995 Oct;104(1):29-38. doi: 10.1007/BF00352223.

Abstract

The ends of eukaryotic chromosomes have special properties and roles in chromosome behavior. Selection for telomere function in yeast, using a Chinese hamster hybrid cell line as the source DNA, generated a stable yeast artificial chromosome clone containing 23 kb of DNA adjacent to (TTAGGG)n, the vertebrate telomeric repeat. The common repetitive element d(GT)n appeared to be responsible for most of the other stable clones. Circular derivatives of the TTAGGG-positive clone that could be propagated in E. coli were constructed. These derivatives identify a single pair of hamster telomeres by fluorescence in situ hybridization. The telomeric repeat tract consists of (TTAGGG)n repeats with minor variations, some of which can be cleaved with the restriction enzyme MnlI. Blot hybridization with genomic hamster DNA under stringent conditions confirms that the TTAGGG tracts are cleaved into small fragments due to the presence of this restriction enzyme site, in contrast to mouse telomeres. Additional blocks of (TTAGGG)n repeats are found approximately 4-5 kb internally on the clone. The terminal region of the clone is dominated by a novel A-T rich 78 bp tandemly repeating sequence; the repeat monomer can be subdivided into halves distinguished by more or less adherence to the consensus sequence. The sequence in genomic DNA has the same tandem organization in probably a single primary locus of >20-30 kb and is thus termed a minisatellite.

摘要

真核生物染色体末端在染色体行为中具有特殊性质和作用。利用中国仓鼠杂交细胞系作为源DNA,在酵母中对端粒功能进行筛选,产生了一个稳定的酵母人工染色体克隆,该克隆包含与脊椎动物端粒重复序列(TTAGGG)n相邻的23 kb DNA。常见的重复元件d(GT)n似乎是大多数其他稳定克隆的原因。构建了可在大肠杆菌中繁殖的TTAGGG阳性克隆的环状衍生物。这些衍生物通过荧光原位杂交鉴定出一对仓鼠端粒。端粒重复序列由(TTAGGG)n重复序列组成,存在微小变异,其中一些可被限制性内切酶MnlI切割。与基因组仓鼠DNA在严格条件下进行印迹杂交证实,与小鼠端粒不同,由于存在该限制性酶切位点,TTAGGG片段被切割成小片段。在克隆内部约4 - 5 kb处还发现了额外的(TTAGGG)n重复序列块。克隆的末端区域由一个新的富含A - T的78 bp串联重复序列主导;重复单体可细分为两半,其区别在于对共有序列的或多或少的遵循。基因组DNA中的序列在可能大于20 - 30 kb的单个主要位点具有相同的串联组织,因此被称为微卫星。

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