Katoh M, Nakagawa Y, Yawata T, Kumano S, Kobayashi E, Kurimasa A, Kugoh H, Oshimura M
Department of Molecular and Cell Genetics, School of Life Sciences, Tottori University, Japan.
Jpn J Hum Genet. 1995 Dec;40(4):307-17. doi: 10.1007/BF01900597.
To obtain cosmid markers and transcribed sequences from a specific chromosome region, a series of radiation-reduced hybrids (RHs) containing various regions of human chromosome 11 was prepared from microcell hybrid A9 (neo11) cells containing a normal human chromosome 11 tagged with pSV2neo at 11p11.2. Among 15 radiation hybrid clones isolated, RH(11)-9 which contains a q23 fragment in addition to the neo integration site, was used for the construction of a cosmid library. Cosmid clones having human DNA sequences were screened, and localized by Southern hybridization with the radiation hybrid panel. Fifty-nine cosmids were assigned to 11q23 and 6 cosmids to 11p11.2. Exon amplification proceeded with 23 of the 59 cosmids and 16 putative exons were cloned. Three of them were identical to those constituting a known gene which locates on q23 (ATDC), and the others were unknown. Thus, the RHs containing various subchromosomal fragments of chromosome 11 were useful for constructing region-specific DNA markers. The RH(11)-9 cells and putative exons also facilitate the positional cloning of genes in the 11q23 region.
为了从特定染色体区域获得黏粒标记和转录序列,从含有在11p11.2处用pSV2neo标记的正常人类11号染色体的微细胞杂种A9(neo11)细胞中制备了一系列包含人类11号染色体不同区域的辐射减少杂种(RH)。在分离出的15个辐射杂种克隆中,除了neo整合位点外还包含q23片段的RH(11)-9用于构建黏粒文库。筛选出具有人类DNA序列的黏粒克隆,并通过与辐射杂种板进行Southern杂交进行定位。59个黏粒被定位到11q23,6个黏粒被定位到11p11.2。对59个黏粒中的23个进行外显子扩增,并克隆了16个推定的外显子。其中3个与构成位于q23的已知基因(ATDC)的外显子相同,其他外显子未知。因此,包含11号染色体各种亚染色体片段的RH对于构建区域特异性DNA标记很有用。RH(11)-9细胞和推定的外显子也有助于11q23区域基因的定位克隆。