本文選題：MEXA基因 + 膀胱癌　； 參考：《中國(guó)醫(yī)科大學(xué)學(xué)報(bào)》2017年12期

【摘要】：目的構(gòu)建針對(duì)MEX3A基因的shRNA慢病毒載體,建立穩(wěn)定干擾MEX3A基因表達(dá)的膀胱癌細(xì)胞株。方法采用實(shí)時(shí)PCR檢測(cè)膀胱癌細(xì)胞株中MEX3A基因的表達(dá);應(yīng)用GV115質(zhì)粒構(gòu)建重組靶向MEX3A基因的shRNA慢病毒載體,鑒定及測(cè)序合格后,與包裝載體共轉(zhuǎn)染293T細(xì)胞產(chǎn)生慢病毒顆粒,病毒滴度測(cè)定后轉(zhuǎn)染膀胱癌細(xì)胞,經(jīng)抗生素篩選建立穩(wěn)定表達(dá)si RNA的細(xì)胞株,實(shí)時(shí)PCR檢測(cè)敲減效率。結(jié)果 MEX3A基因在膀胱癌5637和T24細(xì)胞中均有表達(dá),且5637的表達(dá)量高于T24;鑒定及測(cè)序結(jié)果顯示成功構(gòu)建MEX3A-shRNA慢病毒載體,包裝后病毒滴度較高,實(shí)時(shí)PCR結(jié)果表明慢病毒轉(zhuǎn)染5637細(xì)胞后能穩(wěn)定抑制MEX3A基因的表達(dá)。結(jié)論應(yīng)用慢病毒介導(dǎo)的RNA干擾技術(shù)可成功建立穩(wěn)定抑制MEX3A基因表達(dá)的細(xì)胞株。
[Abstract]:Objective to construct a shRNA lentivirus vector targeting MEX3A gene and to establish bladder cancer cell line which interferes with the expression of MEX3A gene. Methods the expression of MEX3A gene in bladder cancer cell line was detected by real-time PCR, and the recombinant shRNA lentivirus vector targeting MEX3A gene was constructed by using GV115 plasmid. After qualified identification and sequencing, lentivirus particles were co-transfected into 293T cells with packaging vector. After transfection into bladder cancer cells by titer assay, stable expression of si RNA was established by antibiotic screening, and knockout efficiency was detected by real time PCR. Results MEX3A gene was expressed in 5637 and T24 cells of bladder cancer, and the expression of 5637 was higher than that of T24.The identification and sequencing showed that MEX3A-shRNA lentivirus vector was successfully constructed, and the titer of MEX3A-shRNA lentivirus was higher after packaging. The results of real-time PCR showed that lentivirus transfected 5637 cells could stably inhibit the expression of MEX3A gene. Conclusion Lentivirus-mediated RNA interference technique can be successfully used to establish a cell line that stably inhibits the expression of MEX3A gene.
【作者單位】： 中國(guó)醫(yī)科大學(xué)附屬盛京醫(yī)院超聲科;
【基金】：國(guó)家自然科學(xué)基金(81371552)
【分類號(hào)】：R737.14

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