人MAGE-12基因的克隆与序列分析  

Cloning and Sequencing of the Tumor Antigen MAGE-12 Gene Fragment.

作　　者：张国俊[1] 赵国强[2] 王华启[1] 张世杰[2] 王蕾[2] 楚荷莹[1] 

机构地区：[1]郑州大学第一附属医院呼吸内科,450052 [2]郑州大学基础医学院微生物与免疫教研室

出　　处：《医学研究杂志》2006年第3期20-23,共4页Journal of Medical Research

基　　金：河南省医学科技创新人才工程

摘　　要：目的克隆人MAGE-12基因并测序。方法从人肺癌组织中提取mRNA,用RT-PCR从中扩增出MAGE-12基因片段,对其进行PvuⅡ酶切鉴定;将扩增片段克隆于pGEM-Teasy载体中,转化JM109菌;进行蓝白筛选后,运用pGEM-Teasy质粒的T7/SP6作为引物对重组子鉴定,并进行DNA测序。结果PCR扩增得到944bp的基因片段,经PvuⅡ酶切初步证实为MAGE-12基因;阳性克隆经筛选鉴定连接有目的基因;目的基因测序结果与GenBank比较,结果完全相同,表明靶基因成功插入pGEM-Teasy。结论成功克隆MAGE-12基因,为MAGE-12用于肿瘤的生物治疗做出准备工作。Objective To clone the MAGE - 12 gene, preparing for researching its biological effects on MAGE - 12 positive malignant tumors. Methods mRNA was extracted from human lung cancer specimen. MAGE - 12 gene was amplified by reverse transcriptase polymerase chain reaction （RT- PCR） and was identified by enzyme cutting with PvuⅡ .The PCR fragment was inserted into pGEM - T easy plasmid, and then was transformed into JM109. After the selection of blue/white screening, the primers designed with T7/SP6 sequence of pGEM - T easy plasmid were applied to identify the recombinant. Further more, DNA sequence of the recombinant was analyzed. Results The length of the DNA fragment RT - PCR amplified by mRNA was 944 base pairs, which was conformed by enzyme Pvu Ⅱ cutting identification. The correctness of linking between MAGE - 12 and pGEM - T easy was verified through selection of blue/white screening and identified by T7/SP6 primers, and the sequences of DNA fragment were homology with corresponding sequences published in GenBank, which indicated that the target gene had been inserted into pGEM - T easy successfully. Conclusion The MAGE - 12 gene was cloned successfully, which make the foundation of biological treatment by using MAGE - 12 gene.

关 键 词：MAGE-12 肿瘤抗原 基因 

分 类 号：R734.2[医药卫生—肿瘤] R-052[医药卫生—临床医学]