包含1.5倍人类21号染色体的酵母人工染色体文库的构建与表征

Construction and characterization of a yeast artificial chromosome library containing 1.5 equivalents of human chromosome 21.

作者信息

Potier M C, Kuo W L, Dutriaux A, Gray J, Goedert M

机构信息

Medical Research Council Laboratory of Molecular Biology, Cambridge, United Kingdom.

出版信息

Genomics. 1992 Oct;14(2):481-3. doi: 10.1016/s0888-7543(05)80245-9.

DOI:10.1016/s0888-7543(05)80245-9

PMID:1427862

Abstract

A library of yeast artificial chromosomes (YACs) was constructed from a human/hamster somatic cell hybrid containing human chromosome 21 (q11-qter). Cells were embedded in agarose, and the DNA was partially digested with EcoRI, released into solution by agarase treatment of the agarose plugs, ligated into pYAC4, and transferred into yeast. Double screening of the yeast transformants with human and hamster genomic DNA allowed the selection of clones hybridizing only with human DNA. The library consists of 321 clones, amounting to 1.5 equivalents (61 Mb) of chromosome 21. The mean YAC size calculated from 178 clones is 190 +/- 100 kb. Screening of the library with eight sequence-tagged sites gave six positives. Among 21 YACs tested by in situ hybridization, 17 mapped to chromosome 21.

摘要

利用一个含有人类21号染色体（q11 - qter）的人/仓鼠体细胞杂种构建了酵母人工染色体（YAC）文库。将细胞包埋在琼脂糖中，用EcoRI对DNA进行部分消化，通过对琼脂糖凝胶块进行琼脂糖酶处理将DNA释放到溶液中，连接到pYAC4中，然后转入酵母。用人和仓鼠基因组DNA对酵母转化子进行双重筛选，从而选择出仅与人DNA杂交的克隆。该文库由321个克隆组成，相当于21号染色体的1.5倍（61 Mb）。根据178个克隆计算出的平均YAC大小为190±100 kb。用8个序列标签位点筛选该文库得到6个阳性克隆。在通过原位杂交检测的21个YAC中，有17个定位于21号染色体。