Syk基因转染对人结肠癌细胞系影响的实验研究

目的构建Syk基因逆转录病毒载体,并观察其对人结肠癌细胞生物学特性的影响。方法通过RT-PCR扩增出Syk基因的部分cDNA片段,将其插入到逆转录病毒载体pLNCX上,构建成Syk基因的表达载体,并经酶切鉴定和测序确认。将Syk基因表达载体经脂质体介导转染人结肠腺癌细胞系LoVo,筛选出Syk基因稳定表达的细胞株,通过Southern blot检测Syk基因的整合;RT-PCR及DNA测序法检测Syk基因的转录;通过Western blot检测转染细胞Syk基因的表达;二苯基溴化四氮唑蓝(MTT)法检测细胞增殖。结果 Syk基因已整合入细胞并获稳定表达,并显著抑制人结肠腺癌细胞LoVo的增殖。结论 Syk基因逆转录病毒载体通过外源Syk基因mRNA水平的表达,从而抑制人结肠腺癌细胞的生长。

Experimental Study On Syk Gene Transfection To Human Colon Cancer Cell Lines

Objective To construct the retrovirus vector carrying Syk gene,the effect of Syk gene transfection on the growth of human colon cancer cell lines was observed.Methods The cDNA fragment of the Syk gene was PCR-amplified and inserted into the retrovirus vector pLNCX.The constructed Syk expression vector was proved to be the same as designed by restriction endonuclease analysis and sequencing.LoVo cells were transfected with the recombinant retrovirus vector by using Lipofectamine reagent according to the manufacturer＇s instructions.By using Southern blot,RT-PCR amplification,DNA sequencing,MTT,stable cell clones expressing Syk were detected and observed for the changes of their biologic characteristics.Results Exogenous Syk was successfully transferred into LoVo cells and obtained stable expression.The Syk gene could significantly repress the proliferation of LoVo cells.Conclusion Effects of Syk gene on LoVo cell growth was caused by its expression of exogenous Syk mRNA.