天堂国产午夜亚洲专区-少妇人妻综合久久蜜臀-国产成人户外露出视频在线-国产91传媒一区二区三区

當(dāng)前位置:主頁(yè) > 醫(yī)學(xué)論文 > 眼科論文 >

人IL-24基因?qū)D133陽(yáng)性喉癌細(xì)胞作用的實(shí)驗(yàn)研究

發(fā)布時(shí)間:2019-01-19 10:57
【摘要】:目的:克隆人IL-24基因,并構(gòu)建其真核表達(dá)載體;分離Hep-2喉癌細(xì)胞株中CD133陽(yáng)性細(xì)胞(CD133+Hep-2),通過(guò)研究人IL-24基因在CD133+Hep-2細(xì)胞中的表達(dá),探討IL-24基因?qū)D133+Hep-2細(xì)胞的作用。 方法:分離正常人外周血單個(gè)核細(xì)胞,應(yīng)用Trizol法提取單個(gè)核細(xì)胞的總RNA,將總RNA應(yīng)用RT-PCR法反轉(zhuǎn)錄為cDNA,設(shè)計(jì)引物P1、P2,擴(kuò)增人IL-24基因,瓊脂糖凝膠電泳及基因測(cè)序正確后將其TA克隆入pMD19-T simple vector載體;通過(guò)NheI、XhoI雙酶切人IL-24基因和真核表達(dá)載體pIRES2-ZsGreen1,構(gòu)建人IL-24基因的真核表達(dá)載體pIRES2-ZsGreen1-hIL-24并應(yīng)用酶切及基因測(cè)序驗(yàn)證;流式細(xì)胞儀(FCM)從Hep-2細(xì)胞中分選CD133+Hep-2細(xì)胞,將驗(yàn)證正確的pIRES2-ZsGreen1-hIL-24在脂質(zhì)體2000介導(dǎo)下轉(zhuǎn)入CD133+Hep-2細(xì)胞,并驗(yàn)證其表達(dá)情況;MTT、FCM檢測(cè)人IL-24基因?qū)D133+Hep-2細(xì)胞生長(zhǎng)的影響;裸鼠移植瘤實(shí)驗(yàn)檢測(cè)轉(zhuǎn)染人IL-24基因的CD133+Hep-2細(xì)胞的成瘤能力。 結(jié)果:在人外周血單個(gè)核細(xì)胞中成功克隆了621bp的人IL-24基因,基因序列測(cè)定結(jié)果與GenBank中報(bào)道一致;pMD19-T-hIL-24、pIRES2-ZsGreen1-hIL-24重組表達(dá)質(zhì)粒酶切、基因測(cè)序驗(yàn)證正確;用脂質(zhì)體介導(dǎo)法將重組質(zhì)粒pIRES2-ZsGreen1-hIL-24成功轉(zhuǎn)染入CD133+Hep-2細(xì)胞,經(jīng)檢驗(yàn)人IL-24基因整合入轉(zhuǎn)染細(xì)胞并穩(wěn)定表達(dá);MTT法檢測(cè)結(jié)果顯示轉(zhuǎn)入IL-24基因?qū)嶒?yàn)組的細(xì)胞生長(zhǎng)與未轉(zhuǎn)染及空載體對(duì)照組相比明顯受到抑制(P0.05); FCM檢測(cè)細(xì)胞凋亡的結(jié)果顯示實(shí)驗(yàn)組與未轉(zhuǎn)染及空載體對(duì)照組相比凋亡率明顯增加(P0.05);裸鼠移植瘤實(shí)驗(yàn)證實(shí)轉(zhuǎn)入hIL-24基因的實(shí)驗(yàn)組CD133+Hep-2細(xì)胞裸鼠移植瘤的體積、重量明顯低于未轉(zhuǎn)染及空載體對(duì)照組移植瘤(P0.05)。 結(jié)論:人IL-24基因的表達(dá)能夠抑制CD133+Hep-2細(xì)胞的增殖,促進(jìn)其凋亡,,抑制CD133+Hep-2細(xì)胞裸鼠移植瘤的生長(zhǎng)。
[Abstract]:Objective: to clone human IL-24 gene and construct its eukaryotic expression vector. CD133 positive cells (CD133 Hep-2) were isolated from Hep-2 laryngeal carcinoma cell lines. The effect of IL-24 gene on CD133 Hep-2 cells was studied by studying the expression of human IL-24 gene in CD133 Hep-2 cells. Methods: normal human peripheral blood mononuclear cells (PBMC) were isolated. Total RNA, of mononuclear cells was extracted by Trizol method. Total RNA was reverse transcribed into cDNA, designed primer P1CP2to amplify human IL-24 gene. The TA was cloned into pMD19-T simple vector after correct agarose gel electrophoresis and gene sequencing. The eukaryotic expression vector pIRES2-ZsGreen1-hIL-24 of human IL-24 gene was constructed by NheI,XhoI double enzyme digestion of human IL-24 gene and eukaryotic expression vector pIRES2-ZsGreen1, and verified by enzyme digestion and gene sequencing. Flow cytometry (FCM) (FCM) was used to separate CD133 Hep-2 cells from Hep-2 cells. The correct pIRES2-ZsGreen1-hIL-24 was transferred into CD133 Hep-2 cells mediated by liposome 2000 and its expression was confirmed. The effect of human IL-24 gene on the growth of CD133 Hep-2 cells was detected by MTT,FCM, and the tumorigenic ability of CD133 Hep-2 cells transfected with human IL-24 gene was detected by tumor transplantation assay in nude mice. Results: the human IL-24 gene of 621bp was cloned successfully in human peripheral blood mononuclear cells, and the results of gene sequencing were consistent with those reported in GenBank. The recombinant plasmid pIRES2-ZsGreen1-hIL-24 was successfully transfected into CD133 Hep-2 cells by liposome-mediated transfection, and the human IL-24 gene was integrated into the transfected cells and expressed stably. The results of MTT assay showed that the growth of the cells transferred into the IL-24 gene group was significantly inhibited compared with the untransfected and empty vector control group (P0.05). The results of FCM showed that the apoptotic rate of the experimental group was significantly higher than that of the untransfected and empty vector control group (P0.05). Tumor transplantation in nude mice confirmed that the volume and weight of transplanted tumor of CD133 Hep-2 cells transferred to hIL-24 gene in nude mice were significantly lower than those in untransfected and empty vector control group (P0.05). Conclusion: the expression of human IL-24 gene can inhibit the proliferation of CD133 Hep-2 cells, promote their apoptosis and inhibit the growth of xenografts of CD133 Hep-2 cells in nude mice.
【學(xué)位授予單位】:吉林大學(xué)
【學(xué)位級(jí)別】:博士
【學(xué)位授予年份】:2014
【分類號(hào)】:R739.65

【共引文獻(xiàn)】

相關(guān)期刊論文 前10條

1 吳黎明;程彩濤;陳先祥;王江華;劉海燕;田林;;肝細(xì)胞癌組織CD133與VEGF及CD34表達(dá)預(yù)后意義分析[J];中華腫瘤防治雜志;2013年13期

2 王志強(qiáng);;sFRP、WIF-1、CD_(133)、CD_(44)在肺癌組織中的表達(dá)及其臨床意義[J];實(shí)用心腦肺血管病雜志;2014年02期

3 侯紅麗;李莉;王贊宏;曹濤;王真;;IL-24對(duì)宮頸癌細(xì)胞株Siha細(xì)胞生長(zhǎng)和凋亡的影響[J];現(xiàn)代婦產(chǎn)科進(jìn)展;2009年08期

4 程涵蓉;文飛球;李博;陳麗;溫愛(ài)惠;;IL-24體外誘導(dǎo)兒童急性白血病骨髓單個(gè)核細(xì)胞凋亡研究[J];中國(guó)實(shí)驗(yàn)血液學(xué)雜志;2011年01期

5 程涵蓉;文飛球;李博;黃進(jìn)潔;;兒童急性白血病血清白細(xì)胞介素24的測(cè)定及意義[J];中國(guó)小兒血液與腫瘤雜志;2010年06期

6 楊耿兵;何倩;黃青紅;胡徐龐;劉立;錢程;駱菁菁;;雙靶向溶瘤腺病毒聯(lián)合奧沙利鉑對(duì)腫瘤細(xì)胞凋亡的研究[J];浙江理工大學(xué)學(xué)報(bào);2011年05期

7 溫瑩浩;殷正豐;康曉燕;李瑾;錢海華;陳瑋;;腺病毒介導(dǎo)IL-24聯(lián)合化療藥物增強(qiáng)對(duì)肝癌細(xì)胞PLC/PRF/5增殖的抑制[J];中國(guó)腫瘤生物治療雜志;2007年04期

8 潘s

本文編號(hào):2411307


資料下載
論文發(fā)表

本文鏈接:http://sikaile.net/yixuelunwen/yank/2411307.html


Copyright(c)文論論文網(wǎng)All Rights Reserved | 網(wǎng)站地圖 |

版權(quán)申明:資料由用戶37878***提供,本站僅收錄摘要或目錄,作者需要?jiǎng)h除請(qǐng)E-mail郵箱bigeng88@qq.com
中文字幕区自拍偷拍区| 国产乱人伦精品一区二区三区四区| 五月综合婷婷在线伊人| 精品亚洲av一区二区三区| 精品一区二区三区三级视频| 亚洲中文字幕人妻av| 国产精品午夜小视频观看| 经典欧美熟女激情综合网| 欧美精品一区久久精品| 国产日韩欧美一区二区| 年轻女房东2中文字幕| 91日韩在线视频观看| 男女一进一出午夜视频| 91欧美日韩一区人妻少妇| 高清欧美大片免费在线观看| 国产成人国产精品国产三级| 国产欧美日韩精品成人专区| 国产一区二区三区不卡| 99久久精品视频一区二区| 久久国产成人精品国产成人亚洲| 99国产高清不卡视频| 久久精品国产99精品亚洲| 香蕉网尹人综合在线观看| 日本在线视频播放91| 激情丁香激情五月婷婷| 国产精品熟女乱色一区二区| 欧洲亚洲精品自拍偷拍| 久热香蕉精品视频在线播放| 国产高清视频一区不卡| 亚洲精选91福利在线观看 | 99久久精品国产麻豆| 日韩三极片在线免费播放| 日韩一级欧美一级久久| 91老熟妇嗷嗷叫太91| 久草精品视频精品视频精品 | 不卡中文字幕在线视频| 狠狠干狠狠操亚洲综合| 国产精品一区二区视频| 欧美三级精品在线观看| 欧美日韩国产综合在线| 国产精品欧美一级免费|