Jang W, Hua A, Spilson S V, Miller W, Roe B A, Meisler M H
Department of Human Genetics, University of Michigan, Ann Arbor, Michigan 48109-0618, USA.
Genome Res. 1999 Jan;9(1):53-61.
The mnd2 mutation on mouse chromosome 6 produces a progressive neuromuscular disorder. To determine the gene content of the 400-kb mnd2 nonrecombinant region, we sequenced 108 kb of mouse genomic DNA and 92 kb of human genomic sequence from the corresponding region of chromosome 2p13.3. Three genes with the indicated sizes and intergenic distances were identified: D6Mm5e (>/=81 kb)-787 bp-DOK (2 kb)-845 bp-LOR2 (>/=6 kb). D6Mm5e is expressed in many tissues at very low abundance and the predicted 526-residue protein contains no known functional domains. DOK encodes the p62(dok) rasGAP binding protein involved in signal transduction. LOR2 encodes a novel lysyl oxidase-related protein of 757 amino acid residues. We describe a simple search protocol for identification of conserved internal exons in genomic sequence. Evolutionary conservation proved to be a useful criterion for distinguishing between authentic exons and artifactual products obtained by exon amplification, RT-PCR, and 5' RACE. Conserved noncoding sequence elements longer than 80 bp with >/=75% nucleotide sequence identity comprise approximately 1% of the genomic sequence in this region. Comparative analysis of this human and mouse genomic DNA sequence was an efficient method for gene identification and is independent of developmental stage or quantitative level of gene expression. [The sequence data described in this paper have been submitted to the GenBank data library under the following accession numbers: AC003061, mouse BAC clone 245c12; AC003065, human BAC clone h173(E10); AF053368, mouse Lor2 cDNA; AF084363, 108-kb contig from mouse BAC 245c12; AF084364, mouse D6Mm5e cDNA.]
小鼠6号染色体上的mnd2突变会导致一种进行性神经肌肉疾病。为了确定400kb的mnd2非重组区域的基因组成，我们对来自2p13.3染色体相应区域的108kb小鼠基因组DNA和92kb人类基因组序列进行了测序。鉴定出了三个具有所示大小和基因间距离的基因：D6Mm5e（≥81kb）-787bp-DOK（2kb）-845bp-LOR2（≥6kb）。D6Mm5e在许多组织中以非常低的丰度表达，预测的526个残基的蛋白质不包含已知的功能结构域。DOK编码参与信号转导的p62(dok)rasGAP结合蛋白。LOR2编码一种由757个氨基酸残基组成的新型赖氨酰氧化酶相关蛋白。我们描述了一种用于在基因组序列中鉴定保守内部外显子的简单搜索方案。进化保守性被证明是区分真实外显子与通过外显子扩增、RT-PCR和5'RACE获得的人为产物的有用标准。长度超过80bp且核苷酸序列同一性≥75%的保守非编码序列元件约占该区域基因组序列的1%。对该人类和小鼠基因组DNA序列的比较分析是一种有效的基因鉴定方法，且与发育阶段或基因表达的定量水平无关。[本文所述的序列数据已以下列登录号提交至GenBank数据库：AC003061，小鼠BAC克隆245c12；AC003065，人类BAC克隆h173(E10)；AF053368，小鼠Lor2 cDNA；AF084363，来自小鼠BAC 245c12的108kb重叠群；AF084364，小鼠D6Mm5e cDNA。]
