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目的 进一步分析从肺癌多药耐药细胞株 (SPC A 1/CDDP)克隆出的差异表达基因染色体的定位 ,并鉴定其耐药相关功能。方法 登录美国国家生物技术信息中心网 (www ncbi nlm nih gov) ,根据克隆的目的基因序列数据 ,查询人类基因库 ,分析确定该基因在人类染色体上的位置。以DNA重组技术构建该目的基因的反义表达载体 ,电穿孔法将其转染多药耐药细胞株SPC A 1/CDDP ,采用半定量RT PCR研究该基因表达受抑情况 ,并以MTT法分析转染细胞对几种化疗药物的敏感性改变。结果 电子信息学确定该目的基因定位在人类染色体的 19q13 3～19q13 4位点上。成功构建该基因的反义表达载体 ,转染反义表达载体的SPC A 1/CDDP细胞的目的基因mRNA含量明显减少 ,即基因表达受抑。MTT药敏测试发现转染了反义表达载体的SPC A 1/CDDP细胞对顺铂、阿霉素、5 氟尿嘧啶、长春新碱、足叶乙苷和丝裂霉素 6种化疗药物的耐药指数分别是 3 87、3 2 8、6 71、2 6 5、2 11、5 0 2 ;对前五种化疗药物的耐药指数与对照细胞相比分别下降了 2～3倍。表明该基因与肿瘤细胞的化疗敏感性密切相关。结论 该差异表达基因是一条肺癌耐药相关新基因 ,其在人类染色体的位置为 19q13 3～ 19q13 4。
Objective To further analyze the location of differentially expressed genes cloned from lung cancer multidrug resistance cell line (SPC A 1 / CDDP) and to identify their drug resistance-related functions. METHODS: The human gene pool was searched based on the cloned target gene sequence data and analyzed to determine the position of the gene on the human chromosome at www.ncbi nlm nih gov. The antisense expression vector of the target gene was constructed by DNA recombination technology and transfected into multidrug resistant cell line SPC A 1 / CDDP by electroporation. The expression of the gene was inhibited by semi-quantitative RT-PCR and MTT assay The sensitivity of transfected cells to several chemotherapeutic drugs was analyzed. Results Electron informatics determined that the target gene was mapped on the 19q13 3 ~ 19q13 4 locus in human chromosome. The antisense expression vector of this gene was successfully constructed. The mRNA expression of the target gene in SPC A 1 / CDDP cells transfected with the antisense expression vector was significantly reduced, that is, the gene expression was suppressed. MTT drug sensitivity test found that antisense expression vector SPC A 1 / CDDP cells resistant to cisplatin, doxorubicin, 5-fluorouracil, vincristine, etoposide and mitomycin 6 chemotherapeutic drugs The indexes were 3 87,3 2 8,6 71,2 6 5,2 11,5 0 2 respectively. The resistance index to the first five chemotherapeutic drugs decreased by 2 to 3 times compared with the control cells respectively. This gene is closely related to the chemosensitivity of tumor cells. Conclusion The differentially expressed gene is a new gene associated with drug resistance in lung cancer. Its location in human chromosome is 19q13 3 ~ 19q13 4.