CAJ | 학술논문

培养鹌鹑输卵管上皮细胞和鹌鹑胚胎成纤维细胞作为检测平台,将重组后的人血清白蛋白表达载体用脂质体包埋法转染以上两种细胞,通过荧光显微镜观察到报告基因绿色荧光蛋白的表达,结果表明构建的表达载体在成纤维细胞上未见表达;而原代输卵管上皮细胞上有表达.取转染细胞的基因组DNA,设计特异性的检测引物,用PCR筛选法检测了报告基因表达的阳性细胞克隆,SDS-PAGE银染色法确定鹌鹑输卵管上皮细胞分泌蛋白的分子量,结果显示在转染的鹌鹑输卵管上皮细胞培养上清中含有分子量约为68 kDa的HSA.表明构建的表达载体是有效的,而且表达构件已经初步整合到阳性鹌鹑输卵管上皮细胞的染色体中.
배양암순수란관상피세포화암순배태성섬유세포작위검측평태,장중조후적인혈청백단백표체재체용지질체포매법전염이상량충세포,통과형광현미경관찰도보고기인록색형광단백적표체,결과표명구건적표체재체재성섬유세포상미견표체;이원대수란관상피세포상유표체.취전염세포적기인조DNA,설계특이성적검측인물,용PCR사선법검측료보고기인표체적양성세포극륭,SDS-PAGE은염색법학정암순수란관상피세포분비단백적분자량,결과현시재전염적암순수란관상피세포배양상청중함유분자량약위68 kDa적HSA.표명구건적표체재체시유효적,이차표체구건이경초보정합도양성암순수란관상피세포적염색체중.
we constructed the primary oviduct epithelium and quail embryo fibroblasts cell (QEF)of quail were cultured as testing platform. Post-recombinant expression vector of human serum albumin transfected into the two cells using liposome. The expression of report gene EGFP mainly in cytoplasm was observed under fluoroscope. The results showed thatthere was no expression at fibroblasts of the recombination vectors, but it was expressed in oviduct epithelium primary.Genomic DNA of transfected cells were extracted, specific primers were detected,reporter gene-positive cell clones were detect by using PCR method,and molecular weight of Quail oviduct epithelial cell secretory protein were fixed by using SDS-PAGE.The results showed that there were HAS which molecular weight is 68 kDa in transfected quail oviduct epithelial cellculture supernatant,which proved that the constructed expression vector was effective.And the expression of the initial components have been integrated into the quail oviduct epithelial cells positive for the chromosome.