本文選題：宮頸癌 + 微小核糖核酸　； 參考：《山東醫(yī)藥》2017年28期

【摘要】：目的構(gòu)建微小核糖核酸31(miR-31)慢病毒載體質(zhì)粒,并觀察其對(duì)宮頸癌HeLa細(xì)胞增殖和遷移能力的影響。方法以包含miR-31基因的序列為目的片段,設(shè)計(jì)末端含有相應(yīng)酶切位點(diǎn)的引物,PCR擴(kuò)增目的片段,產(chǎn)物雙酶切后插入線性化慢病毒載體中,構(gòu)建miR-31慢病毒載體質(zhì)粒并鑒定。將miR-31慢病毒載體質(zhì)粒與包裝質(zhì)粒pPACKH1-GAG、pPACKH1-REV、pVSV-G共轉(zhuǎn)染宮頸癌HeLa細(xì)胞,并進(jìn)行慢病毒包裝,檢測(cè)miR-31慢病毒載體質(zhì)粒滴度。取對(duì)數(shù)生長(zhǎng)期HeLa細(xì)胞,隨機(jī)分為空白對(duì)照組、miR-31組、空載體組,空白對(duì)照組不做任何處理,miR-31組予miR-31慢病毒載體質(zhì)粒處理,空載體組予空載體慢病毒質(zhì)粒處理。采用實(shí)時(shí)熒光定量PCR法檢測(cè)各組miR-31 mRNA表達(dá);MTT法和細(xì)胞克隆實(shí)驗(yàn)檢測(cè)各組細(xì)胞增殖能力,Transwell小室法檢測(cè)各組細(xì)胞遷移能力。結(jié)果成功構(gòu)建重組慢病毒表達(dá)質(zhì)粒,其病毒滴度為3×107TU/m L。miR-31組miR-31 mRNA相對(duì)表達(dá)量明顯高于空白對(duì)照組、空載體組(P均0.05),而空白對(duì)照組與空載體組比較P0.05。miR-31組細(xì)胞增殖和遷移能力均高于空白對(duì)照組、空載體組(P均0.05),而空白對(duì)照組與空載體組比較P均0.05。結(jié)論成功構(gòu)建了miR-31慢病毒載體質(zhì)粒,建立了穩(wěn)定過(guò)表達(dá)miR-31的宮頸癌HeLa細(xì)胞。穩(wěn)定過(guò)表達(dá)miR-31的宮頸癌HeLa細(xì)胞增殖和遷移能力明顯增強(qiáng)。
[Abstract]:Objective to construct the vector plasmid of minimal ribonucleic acid 31 (miR-31) lentivirus and to observe the effect of miR-31 on the proliferation and migration of cervical cancer HeLa cells. Methods the sequence containing miR-31 gene was used as the target fragment. Primers with corresponding endonuclease sites were designed to amplify the target fragment. The product was digested by double enzyme and inserted into the linearized lentivirus vector to construct and identify the vector of miR-31 lentivirus. The miR-31 lentivirus vector plasmid and the packaging plasmid pPACKH1-GAGN pPACKH1-REVSV-G were co-transfected into cervical cancer HeLa cells and packaged with lentivirus to detect the titer of miR-31 lentivirus vector plasmid. HeLa cells in logarithmic growth phase were randomly divided into blank control group and empty vector group. The blank control group was treated with miR-31 lentivirus plasmid and empty vector group was treated with empty vector lentivirus plasmid. The expression of miR-31 mRNA in each group was detected by real-time fluorescence quantitative PCR and cell clone assay was used to detect the cell proliferation ability and the migration ability of each group was detected by Transwell chamber assay. Results Recombinant lentivirus expression plasmid was successfully constructed. The relative expression of miR-31 mRNA in the virus titer of 3 脳 10 7 TU / m L.miR-31 group was significantly higher than that in the control group. The cell proliferation and migration ability of P0.05.miR-31 group was higher than that of blank control group (P 0.05), while that of blank control group was 0.05 compared with empty carrier group (P 0.05). Conclusion the vector plasmid of miR-31 lentivirus was successfully constructed and HeLa cells stably expressed miR-31 were established. The proliferation and migration of HeLa cells with stable expression of miR-31 were significantly enhanced.
【作者單位】： 廣西壯族自治區(qū)人民醫(yī)院;
【基金】：國(guó)家自然科學(xué)基金資助項(xiàng)目(81060219、81660434) 廣西自然科學(xué)基金資助項(xiàng)目(2014GXNSFAA118266)
【分類號(hào)】：R737.33

【參考文獻(xiàn)】

相關(guān)期刊論文 前1條

1 孟凡榮;陳琛;萬(wàn)海粟;周清華;;慢病毒載體及其研究進(jìn)展[J];中國(guó)肺癌雜志;2014年12期

【共引文獻(xiàn)】

相關(guān)期刊論文 前10條

1 王琳琳;胡曉霞;劉斐;;miR-31慢病毒載體質(zhì)粒構(gòu)建及其對(duì)宮頸癌HeLa細(xì)胞增殖和遷移能力的影響[J];山東醫(yī)藥;2017年28期

2 鄒斌;周學(xué)亮;詹宇亮;陳紫晴;賴松青;吳霞;劉季春;;慢病毒載體介導(dǎo)HIF1α穩(wěn)定表達(dá)A549細(xì)胞系構(gòu)建[J];重慶醫(yī)學(xué);2017年20期

3 常海平;楊彩榮;鄭健;;人TPX2基因過(guò)表達(dá)慢病毒質(zhì)粒的構(gòu)建及其人宮頸癌細(xì)胞穩(wěn)定表達(dá)株的篩選[J];中國(guó)生物制品學(xué)雜志;2017年06期

4 張濤;王攀;馬珊珊;石振慶;程康;黃團(tuán)結(jié);劉艷霞;楊波;關(guān)方霞;;慢病毒介導(dǎo)的FOXQ1過(guò)表達(dá)對(duì)人臍帶間充質(zhì)干細(xì)胞衰老的影響[J];鄭州大學(xué)學(xué)報(bào)(醫(yī)學(xué)版);2017年03期

5 王子雄;王勤章;錢彪;;經(jīng)尿道灌注SCL基因重組慢病毒的DCP豚鼠膀胱Cajal樣間質(zhì)細(xì)胞數(shù)量、分布及超微結(jié)構(gòu)觀察[J];山東醫(yī)藥;2017年15期

6 張銀花;李秋明;孫英健;沈紅;;NTE基因ShRNA慢病毒載體的構(gòu)建與鑒定[J];北京農(nóng)學(xué)院學(xué)報(bào);2017年02期

7 湯晗;聶建云;李文斌;董鳳萍;;慢病毒載體系統(tǒng)的發(fā)展及在腫瘤基因治療中的應(yīng)用[J];現(xiàn)代腫瘤醫(yī)學(xué);2017年10期

8 汪曉;孫濤;;尾側(cè)型同源轉(zhuǎn)錄因子2慢病毒載體的構(gòu)建及其轉(zhuǎn)染骨髓間充質(zhì)干細(xì)胞的實(shí)驗(yàn)研究[J];解放軍醫(yī)學(xué)院學(xué)報(bào);2017年03期

9 武江波;蘇芳;李齡;朱小東;陳龍;曲頌;莫柒艷;林歡;;穩(wěn)定過(guò)表達(dá)XAF1基因鼻咽癌放射抗拒CNE-2R細(xì)胞株的構(gòu)建[J];中國(guó)癌癥防治雜志;2016年06期

10 魏艷青;馬路平;王江平;錢彪;王勤章;;經(jīng)尿道灌注干細(xì)胞白血病基因慢病毒轉(zhuǎn)染豚鼠糖尿病膀胱病變的效果研究[J];中國(guó)全科醫(yī)學(xué);2016年33期

【二級(jí)參考文獻(xiàn)】

相關(guān)期刊論文 前3條

1 王露;翟瑋瑋;楊向榮;王芳;李見(jiàn);張強(qiáng);李玉云;;慢病毒介導(dǎo)RNA干擾沉默F(xiàn)as基因在臍帶間充質(zhì)干細(xì)胞中的表達(dá)[J];南方醫(yī)科大學(xué)學(xué)報(bào);2014年10期

2 張磊;劉慶友;胡天;張曉溪;崔奎青;陸鳳花;石德順;;第三代慢病毒高效率包裝系統(tǒng)的建立[J];基因組學(xué)與應(yīng)用生物學(xué);2009年02期

3 鄧?yán)^先,沈偉;用慢病毒載體制備轉(zhuǎn)基因動(dòng)物的研究進(jìn)展[J];中國(guó)生物工程雜志;2004年09期

，

本文編號(hào)：2050429