生物信息学技术克隆人类神经髓鞘蛋白零家族基因  

Cloning of Human Myelin Protein Zero-like Genes by Bioinformatics Strategy

作　　者：唐冬生[1] 禹宽平[1] 张华莉[1] 潘乾[1] 戴和平[1] 夏家辉[1] 

机构地区：[1]湖南医科大学中国医学遗传学国家重点实验室,长沙410078

出　　处：《生物化学与生物物理学报》2000年第4期364-368,共5页

基　　金：国家重点基础研究发展规划项目 !("973");"863"计划 !(Z19 0 2 0 2 0 2 );国家自然科学基金资助项目&&

摘　　要：为克隆新的髓鞘蛋白相关基因 ,将MPZ基因编码区cDNA序列与EST数据库进行同源性比较 ,得到与MPZ显著相似的 2个EST ,构建成 80 1bp的重叠群。此重叠群与定位在 1q2 4的 12 8kbgDNA的相似性为 10 0 %。计算机分析表明 80 1bp的重叠群可能存在一个 435bp的阅读框架。在重叠群上设计两个引物与文库载体臂上的引物配对 ,扩增各种cDNA文库DNA作巢式PCR。在所得cDNA上再设计引物进行巢式PCR ,最终克隆了人髓鞘蛋白零样基因I型和II型 (MPZL1a、MPZL1b ,GenBank :AF0 95 72 7、AF0 92 42 4)。对MPZL与MPZ的基因结构和推导的蛋白质一级结构进行分析与比较 ,证明MPZL1是MPZ家族的第二个成员。对 2 4个腓骨肌萎缩症家系和 2 6个非综合征型耳聋家系进行了MPZL1基因的突变检测 。To clone novel myelin protein related genes, two human ESTs, which shared significant similarity with the human myelin protein zero gene, were found by the comparison of homologue between the cDNA coding region sequences of MPZ gene and the EST database of NCBI. An 801 bp EST contig was assembled, which was 100% identical with a 128 kb genomic sequence, mapped to 1q24. A 435 bp open reading frame (ORF) within the 801 bp contig was shown by computer analysis. Two primers, designed according to the sequence of the contig, were coupled with the primers(λgt10 5 and gt10 5) on the sequences flanking cloning site of the cDNA library vector to amplify the cDNA library sequences by nested PCR. New primers, designed based on novel cDNA sequences, were used for the PCR amplification with λgt10 5 and gt10 5 in the same way as above. Finally, the human myelin protein zero like gene isoform I and II ( MPZL 1a, MPZL 1b; GenBank: AF095727, AF092424) were cloned. Comparison of gene and protein structures between MPZL1 and MPZ revealed that MPZL 1 is the second member of MPZ family. Mutation analysis of MPZL1 gene was performed in 24 Charcot Marie Tooth disease (CMT) families and 26 nonsyndrome deafness families, but no mutation was found.

关 键 词：髓鞘蛋白零样基因 计算机分析 基因克隆 髓鞘病 

分 类 号：Q785[生物学—分子生物学]