ASPP1对胃癌细胞SGC-7901增殖抑制

Inhibitive effects of ASPP1 genes on the proliferation of gastric cancer cell line SGC-7901

目的本研究扩增及鉴定包含人类ASPP1基因转录本蛋白编码区的cDNA片段,将此片段克隆入真核表达载体,转染胃癌细胞,观察其对胃癌细胞增殖力的影响。方法从胎盘组织中提取总RNA,采用RT-PCR技术扩增ASPP1基因全长编码序列CDS(full-length coding sequence),目的片段回收纯化后用限制性内切酶XbaⅠ,EcoR I酶切的方法鉴定目的片段正确与否并检测其纯度、浓度;并转染胃癌SGC-7901细胞,观察ASPP1蛋白的表达及用MTT法进行细胞增殖力检测。结果该扩增片段经限制性核酸内切酶切图谱分析,与预期结果吻合,p53表达增加,转染的胃癌SGC-7901细胞出现增殖受抑。结论人类ASPP1基因对胃癌细胞增殖有抑制作用,从而为临床上胃癌的基因治疗提供了新思路。

Objective To amplify and identify gene transcripts containing the human ASPP1 protien coding region cDNA fragment,transfect gastric cancer cell line,and observe biological character changes of the trans-fected gastric cancer cells.Methods Using TrizolTM Reagent,total RNA was extracted from human cell,design-ing the particular primer with restriction enzyme XbaⅠ,EcoR I recognition site in its end.ASPP1 gene CDS fragment was amplified by reverse transcription-polymerase chain reaction（RT-PCR）,then the ASPP1cDNA tar-get fragment was purified,and was identified by restriction enzyme digestion analysis.The pcDNA3.1-ASPP1 and pcDNA3.1 were transfected into SGC-7901 cell line by LipofectAMINE 2000.Observing the expression of ASPP1 protien in every class.The cell proliferation was detected by MTT.Results Making use of RT-PCR method and particular primer which include restriction enzyme XbaⅠ,EcoR I recognition enzyme site in its end,this result was consistant with the sequence designed in advance too.The relative expression of p53 in SGC-7901-pcDNA3.1-ASPP1 was significantly higher than those in control SGC-7901 and pcDNA3.1-ASPP1.The proliferative ability of SGC-7901-pcDNA3.1-ASPP1 was much more inhibited than that in the controls.Conclusions ASPP1 gene is successfully amplified,and ASPP1 gene has the inhibitive effect on the proliferation of gastric cancer cell,so ASPP1 is a potential gene therapy strategy for gastric cancer.