CAJ | 학술논문

目的本文旨在构建11β-羟基类固醇脱氢酶2 (11β-HSD2)基因的重组真核表达载体,并检测其转染入哺乳动物细胞后对白介素6(interleukin-6,IL-6)表达水平的影响.方法用PCR方法扩增11β-HSD2基因的全长cDNA,先亚克隆至pGMT载体,测序正确后再亚克隆至pcDNA3.1 myc-hisC 以构建其重组真核表达载体.将重组的真核表达载体瞬时转染人肺腺癌细胞系A549,用Western blot 方法鉴定表达情况,并用ELISA方法检测IL-6表达水平.结果测序表明11β-HSD2的重组真核表达载体构建成功,Western blot显示真核表达载体能够在A549细胞中成功表达,并可使IL-6表达水平升高.结论成功构建了11β-HSD2基因的重组真核表达载体,初步验证了该基因的促炎效应.
목적 본문지재구건11β-간기류고순탈경매2 (11β-HSD2)기인적중조진핵표체재체,병검측기전염입포유동물세포후대백개소6(interleukin-6,IL-6)표체수평적영향.방법 용PCR방법확증11β-HSD2기인적전장cDNA,선아극륭지pGMT재체,측서정학후재아극륭지pcDNA3.1 myc-hisC 이구건기중조진핵표체재체.장중조적진핵표체재체순시전염인폐선암세포계A549,용Western blot 방법감정표체정황,병용ELISA방법검측IL-6표체수평.결과 측서표명11β-HSD2적중조진핵표체재체구건성공,Western blot현시진핵표체재체능구재A549세포중성공표체,병가사IL-6표체수평승고.결론성공구건료11β-HSD2기인적중조진핵표체재체,초보험증료해기인적촉염효응.
Objective To establish recombinant eukaryotic vector of 11β-hydroxysteroid dehydrogenase 2 gene in mammalian cell and to detect the expression of interleukin-6 after the vector transfection.Methods The cDNA of 11β-hydroxysteroid dehydrogenase 2 gene was amplified by PCR.After subcloned with pGMT vector and sequenced,the gene was inserted into the eukaryotic vector pcDNA3.1myc hisC.The expression vector was transiently transfected in human lung adenocarcinoma cell line A549 cells,and then the protein was detected using Western blot,interleukin-6 was measured by ELISA.Results Sequencing revealed 11β-hydroxysteroid dehydrogenase 2 recombinant eukaryotic expression vector was constructed successfully,and Western blot indicated that eukaryotic expression vector expressed successfully in A549 cells.Transfection with recombinant eukaryotic vector of 11β hydroxysteroid dehydrogenase 2 gene significantly increased interleukin 6 expression.Conclusions The recombinant eukaryotic vector of 11β-hydroxysteroid dehydrogenase 2 gene is established successfully and could be highly overexpressed in A549 cells.The increase level of interleukin-6 suggests the proinflammatory effect of 11β-hydroxysteroid dehydrogenase 2.