中华实验外科杂志
中華實驗外科雜誌
중화실험외과잡지
CHINESE JOURNAL OF EXPERIMENTAL SURGERY
2009年
7期
884-886
,共3页
刘卫仁%于颖彦%倪培华%吴建林%计骏%张佳年%陈雪华%刘炳亚%朱正纲
劉衛仁%于穎彥%倪培華%吳建林%計駿%張佳年%陳雪華%劉炳亞%硃正綱
류위인%우영언%예배화%오건림%계준%장가년%진설화%류병아%주정강
胃癌%IRX1%真核表达载体
胃癌%IRX1%真覈錶達載體
위암%IRX1%진핵표체재체
Gastric carcinoma%IRX1%Eukaryotic expression vector
目的 构建pEGFP-IRX1真核表达载体,体外转导至SGC-7901胃癌细胞株,观察IRX1基因转染后蛋白表达与细胞内定位.方法 PCR扩增IRX1全长1443 bp编码序列,将PCR扩增产物连接人pGEM-Teasy载体,经测序确认序列无误后,亚克隆入pEGFP-N1载体,构建pEG-FP-IRX1真核表达载体.采用Lipofectamine 2000转染SGC-7901胃癌细胞株,在荧光显微镜下观察外源性IRX1基因导入后在胃癌细胞内的表达与亚细胞定位.结果 成功构建有绿色荧光蛋白融合的IRX1真核表达载体,外源性IRX1基因在胃癌SGC-7901细胞中表达成功,在荧光显微镜下IRX1表达蛋白定位于细胞核内.结论 pEGFP-IRX1真核表达载体构建成功,并可在细胞内表达,这将为IRX1在胃癌内的功能研究奠定基础.
目的 構建pEGFP-IRX1真覈錶達載體,體外轉導至SGC-7901胃癌細胞株,觀察IRX1基因轉染後蛋白錶達與細胞內定位.方法 PCR擴增IRX1全長1443 bp編碼序列,將PCR擴增產物連接人pGEM-Teasy載體,經測序確認序列無誤後,亞剋隆入pEGFP-N1載體,構建pEG-FP-IRX1真覈錶達載體.採用Lipofectamine 2000轉染SGC-7901胃癌細胞株,在熒光顯微鏡下觀察外源性IRX1基因導入後在胃癌細胞內的錶達與亞細胞定位.結果 成功構建有綠色熒光蛋白融閤的IRX1真覈錶達載體,外源性IRX1基因在胃癌SGC-7901細胞中錶達成功,在熒光顯微鏡下IRX1錶達蛋白定位于細胞覈內.結論 pEGFP-IRX1真覈錶達載體構建成功,併可在細胞內錶達,這將為IRX1在胃癌內的功能研究奠定基礎.
목적 구건pEGFP-IRX1진핵표체재체,체외전도지SGC-7901위암세포주,관찰IRX1기인전염후단백표체여세포내정위.방법 PCR확증IRX1전장1443 bp편마서렬,장PCR확증산물련접인pGEM-Teasy재체,경측서학인서렬무오후,아극륭입pEGFP-N1재체,구건pEG-FP-IRX1진핵표체재체.채용Lipofectamine 2000전염SGC-7901위암세포주,재형광현미경하관찰외원성IRX1기인도입후재위암세포내적표체여아세포정위.결과 성공구건유록색형광단백융합적IRX1진핵표체재체,외원성IRX1기인재위암SGC-7901세포중표체성공,재형광현미경하IRX1표체단백정위우세포핵내.결론 pEGFP-IRX1진핵표체재체구건성공,병가재세포내표체,저장위IRX1재위암내적공능연구전정기출.
Objective To construct the eukaryotic expression vector of pEGFP-IRX1, and observe the IRX1 protein expression and its sub-cellular localization in SGC-7901 gastric cancer cell line. Methods The coding sequence of IRX1 1443 bp was obtained by PCR amplification,then the PCR product was in-serted into the pGEM-Teasy vector. After confirming the DNA sequence, IRX1 amplification product was sub-cloned into pEGFP-N1 vector. Finally, an IRX1 gene eukaryotic expression vector was obtained. The pEGFP-IRX1 expression vector was transfected into SGC-7901 gastric cancer cell line by Lipofectamine 2000,and the expression of pEGFP-IRX1 was detected by GFP fluorescence. Results The recombinant vector was highly expressed in SGC-7901 cells. The green fluorescence of EGFP-1RX1 fusion protein was clearly localized in nuclei of gastric cancer cells. Conclusion pEGFP-IRX1 expression vector was con-structed successfully, and the IRX1 protein was mainly observed in nuclei,which settled a basis for further functional study of gene IRX1 in gastric cancer.
