【摘要】：目的建立穩(wěn)定表達(dá)葉酸受體α(FRα)的C26細(xì)胞,用于FRα基因疫苗的后續(xù)研究。方法采用脂質(zhì)體轉(zhuǎn)染法,將前期構(gòu)建的重組真核表達(dá)質(zhì)粒pc DNA3.1-FRα基因?qū)隒26細(xì)胞,用G418加壓進(jìn)行篩選,并通過單克隆操作,建立穩(wěn)定轉(zhuǎn)染葉酸受體α基因的單克隆細(xì)胞株。利用反轉(zhuǎn)錄PCR及免疫熒光細(xì)胞化學(xué)染色檢測穩(wěn)定轉(zhuǎn)染細(xì)胞中葉酸受體α基因的表達(dá)情況,并利用反轉(zhuǎn)錄PCR檢測傳代細(xì)胞中FRα基因表達(dá)的穩(wěn)定性。結(jié)果經(jīng)轉(zhuǎn)染、G418篩選以及單克隆化操作,得到單克隆FRα基因穩(wěn)定轉(zhuǎn)染細(xì)胞株,所得單克隆穩(wěn)定轉(zhuǎn)染細(xì)胞株可表達(dá)FRαmRNA及蛋白,并能夠在多次傳代后保持表達(dá)的穩(wěn)定性。結(jié)論成功構(gòu)建了FRα穩(wěn)定轉(zhuǎn)染細(xì)胞株。
[Abstract]:Objective to establish C26 cells stably expressing folate receptor 偽 (FR 偽) for further study of FR 偽 gene vaccine. Methods Recombinant eukaryotic expression plasmid pc DNA3.1-FR 偽 was transfected into C26 cells by liposome transfection. A monoclonal cell line stably transfected with folate receptor 偽 gene was established. Reverse transcription PCR and immunofluorescence cytochemical staining were used to detect the expression of folate receptor 偽 gene in stable transfected cells, and reverse transcription PCR was used to detect the stability of FR 偽 gene expression. Results after transfection, G418 screening and monoclonal manipulation, the stable transfection cell line of FR 偽 gene was obtained, and the stable expression of FR 偽 mRNA and protein could be maintained after multiple passages. Conclusion the stable transfection cell line of FR 偽 was successfully constructed.
【作者單位】： 中國藥科大學(xué)生命科學(xué)與技術(shù)學(xué)院分子生物學(xué)教研室;南京中醫(yī)藥大學(xué)中藥學(xué)一級學(xué)科;
【基金】：國家自然科學(xué)基金(81301902,81102899)
【分類號】：R392

【參考文獻(xiàn)】

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