Torigoe K, Harada T, Kusaba H, Uchiumi T, Kohno K, Green E D, Scherer S W, Tsui L C, Schlessinger D, Kuwano M, Wada M
Department of Biochemistry, Kyushu University School of Medicine, Fukuoka, Japan.
Genomics. 1998 Apr 1;49(1):14-22. doi: 10.1006/geno.1997.5200.
A contig of 21 nonchimeric yeast artificial chromosomes (YACs) was previously assembled across 1.5 Mb of the multidrug resistance (MDR) gene (PGY1 and PGY3) region of human chromosome 7q21.1. This region of the human genome has now been subjected to exon amplification to detect the presence of additional genes. Exon trapping was performed directly on the YACs. Sixty-seven gene fragments were isolated and characterized by sequence analysis and comparison with public databases. The localization of these exons in the 1.5-Mb region was determined by hybridization to YAC clones, and they were localized in 11 subregions of YAC contigs. The exon collection includes 21 exons that were identical to known cDNA sequences of PGY1, PGY3, sorcin (SRI), the cDNA similar to the delta subunit of the human amiloride-sensitive Na- channel (SCNED), and 4 cDNAs with unknown function; 43 exons that showed homology/similarity to known cDNA sequences of mouse DMP1, rat COT, mouse and human NADHD, human MDC, 3 cDNAs encoding possible membrane proteins, and 21 other cDNAs; and 3 exons that shared no homology/similarity with any sequence in public databases. The nucleotide sequences of all the PGY1 and PGY3 exons were identical to the corresponding cDNA sequences previously determined, and these exons were localized to the expected positions on the appropriate YAC clones. No other member of the MDR gene family thus appeared to be present in the 1.5-Mb region. The integrated physical and exon maps should prove valuable for both fine mapping and determination of a complete gene map of this segment of the genome.
先前构建了一个由21个非嵌合酵母人工染色体(YAC)组成的重叠群,跨越人类7号染色体q21.1的多药耐药(MDR)基因(PGY1和PGY3)区域的1.5 Mb。人类基因组的这一区域现已进行外显子扩增,以检测其他基因的存在。直接在YAC上进行外显子捕获。通过序列分析并与公共数据库比较,分离并鉴定了67个基因片段。这些外显子在1.5 Mb区域中的定位通过与YAC克隆杂交来确定,它们定位在YAC重叠群的11个子区域中。外显子集合包括21个与PGY1、PGY3、亲环蛋白(SRI)、与人氨氯地平敏感钠通道δ亚基相似的cDNA(SCNED)的已知cDNA序列相同的外显子,以及4个功能未知的cDNA;43个与小鼠DMP1、大鼠COT、小鼠和人类NADHD、人类MDC的已知cDNA序列具有同源性/相似性的外显子,3个编码可能膜蛋白的cDNA,以及21个其他cDNA;还有3个与公共数据库中的任何序列均无同源性/相似性的外显子。所有PGY1和PGY3外显子的核苷酸序列与先前确定的相应cDNA序列相同,并且这些外显子定位在适当YAC克隆上的预期位置。因此,在1.5 Mb区域中似乎不存在MDR基因家族的其他成员。整合的物理图谱和外显子图谱对于该基因组片段的精细定位和完整基因图谱的确定都应具有重要价值。