人M961全长cDNA及剪接异构体的分离与克隆

The Identification and Cloning of Human M961Full-length cDNA and Its Splicing Isoform

目的分离、克隆人M96基因,并初步探索其在不同血细胞系中的表达。方法根据与鼠M96基因有较高同源性的人表达序列标签(EST)设计筛库引物,筛选成人睾丸及胎脑cDNA文库,采用BLAST及CLUSTALW等程序比较分析筛选到的阳性克隆;应用多细胞系杂交膜分析该基因在各细胞系的表达;用氯化血红素诱导K562细胞向红系分化,探讨该基因在此过程中的表达变化。结果从文库中筛出长度分别为1878和1382bp的M96cDNA克隆,二者为剪接异构体,其开放阅读框分别编码491及262个氨基酸,分别命名为M961和M962(GenBank接受号分别为AF072814和AF073293)。Northern杂交结果表明,在所检测的10种血细胞系中,人淋巴细胞白血病细胞系CEM的M961基因表达量最高,在有巨核及红系特点的Hel、Dami及K562白血病细胞系中M961的表达量也较高。在K562细胞分化过程中,M961基因表达略有升高。结论M961基因在某些白血病细胞系中表达量较高。

Objective To identify and clone the gene encoding human M96gene and study its expression spectrum in several blood cell lines.Methods According to the sequence of human EST which was highly homologous to the mouse M96gene,primers used for library screening were synthesized,then the human adult testis and fetal brain cDNA library were screened.The gene was analyzed by making use of BLAST and CLUSTAL W,and its expression spectrum was studied by multiple-cell lines Northern blot analysis.The expression change of M961in cell differentiation was observed by use of K562cell line induced by hemin.Results Two cDNA clones encoding human M96gene were isolated,identified and named as M961,and M962.They were found to be isoforms of each other.Northern blot showed that M961gene was expressed highly in CEM,Hel,Dami and K562cell lines.However,during K562cell line differentiation,process the expression of M961elevated only slightly.Conclusions M961gene was expressed highly in pluripotent cell lines with erythrocytic and megak aryocytic potentials.