本文選題：CCDC8基因 + 乳腺癌細(xì)胞MDA-MB-231��； 參考：《昆明醫(yī)科大學(xué)》2017年碩士論文

【摘要】：[目的]近年來乳腺癌有年輕化的趨勢,而其中三陰性乳腺癌預(yù)后較差。三陰性乳腺癌被認(rèn)為是多階段、多因素、多基因共同作用的結(jié)果。本課題選用三陰性乳腺癌細(xì)胞株MIDA-MB-231。通過慢病毒載體法沉默該細(xì)胞株的CCDC8基因并構(gòu)建裸鼠乳腺癌移植瘤動(dòng)物模型,觀察CCDC8基因沉默后對(duì)乳腺癌MDA-MB-231細(xì)胞增殖能力的影響及對(duì)裸鼠乳腺癌移植瘤生長的影響。[方法]1、以KD1、KD2序列為作用靶點(diǎn)合成兩條siRNA干擾片段并用慢病毒載體法穩(wěn)定轉(zhuǎn)染MDA-MB-231細(xì)胞。2、通過qRT-PCR篩選CCDC8基因敲減率高的慢病毒穩(wěn)定轉(zhuǎn)染細(xì)胞組。3、通過細(xì)胞克隆形成實(shí)驗(yàn)觀察慢病毒穩(wěn)定轉(zhuǎn)染細(xì)胞組的MDA-MB-231細(xì)胞克隆形成能力。4、采用流式細(xì)胞術(shù)觀察慢病毒穩(wěn)定轉(zhuǎn)染細(xì)胞組的MDA-MB-231細(xì)胞周期、凋亡率變化。5、采用MTT檢測慢病毒穩(wěn)定轉(zhuǎn)染細(xì)胞組的MDA-MB-231細(xì)胞增殖能力。6、構(gòu)建裸鼠乳腺癌移植瘤動(dòng)物模型,觀察瘤體體積及重量變化。[結(jié)果]1、qRT-PCR檢測顯示KD2-siRNA能有效抑制CCDC8基因表達(dá)(P0.01)。2、細(xì)胞克隆形成實(shí)驗(yàn)結(jié)果顯示MDA-MB-231細(xì)胞經(jīng)CCDC8基因沉默后細(xì)胞克隆形成能力降低(P 0.01)。3、流式細(xì)胞術(shù)檢測結(jié)果顯示MDA-MB-231細(xì)胞經(jīng)CCDC8基因沉默后細(xì)胞周期阻滯于G1期(P0.01)且凋亡率增加(P0.01)。4、MTT檢測顯示MDA-MB-231細(xì)胞經(jīng)CCDC8基因沉默后細(xì)胞增殖能力受到抑制(P 0.01)。5、裸鼠皮下注入CCDC8基因沉默后的MDA-MB-231細(xì)胞后,裸鼠乳腺癌移植瘤的生長減緩(P 0.01)。[結(jié)論]1.三陰性乳腺癌細(xì)胞MDA-MB-231經(jīng)CCDC8基因沉默后其細(xì)胞增殖能力減低、細(xì)胞凋亡率增加。2. CCDC8基因沉默可以抑制裸鼠乳腺癌移植瘤的生長。
[Abstract]:Objective: breast cancer tends to be younger in recent years, but the prognosis of three negative breast cancer is poor.Triple-negative breast cancer is considered to be the result of multi-stage, multi-factor and multi-gene interaction.In this study, three negative breast cancer cell line MIDA-MB-231 was selected.The CCDC8 gene of the cell line was silenced by lentivirus vector method and an animal model of breast cancer transplantation in nude mice was established. The effects of CCDC8 gene silencing on the proliferation of breast cancer MDA-MB-231 cells and the growth of breast cancer transplanted tumor in nude mice were observed.[methods] 1. Two siRNA interference fragments were synthesized with KD1hkD2 sequence as the target and stably transfected into MDA-MB-231 cells by lentivirus vector method. The stable transfection cell group of lentivirus with high CCDC8 knockout rate was screened by qRT-PCR, and the cell clone was formed.The clone forming ability of MDA-MB-231 cells in lentivirus stable transfection group was observed. Flow cytometry was used to observe the MDA-MB-231 cell cycle in the lentivirus stable transfected cell group.MTT was used to detect the proliferative ability of MDA-MB-231 cells in lentivirus stable transfection group. The nude mice model of breast cancer transplantation was established and the changes of tumor volume and weight were observed.[results] 1qRT-PCR assay showed that KD2-siRNA could effectively inhibit the expression of CCDC8 gene, and the result of cell clone formation experiment showed that the cell clone formation ability of MDA-MB-231 cells silenced by CCDC8 gene was decreased by CCDC8 gene, and that of MDA-MB-231 cells by CCDC8 was decreased by flow cytometry.After gene silencing, cell cycle arrest in G1 phase P0.01) and apoptosis rate increased P0.01. 4MTT assay showed that the proliferation of MDA-MB-231 cells was inhibited by CCDC8 gene silencing. After subcutaneous injection of CCDC8 gene silenced MDA-MB-231 cells in nude mice, the cell proliferation was inhibited.The growth of breast cancer xenografts in nude mice was slowed down (P 0.01).[conclusion] 1.Three negative breast cancer cell line MDA-MB-231 was silenced by CCDC8 gene. The cell proliferation ability was decreased and the apoptosis rate was increased by 2. 2.CCDC8 gene silencing can inhibit the growth of breast cancer xenografts in nude mice.
【學(xué)位授予單位】：昆明醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R737.9

【參考文獻(xiàn)】

相關(guān)期刊論文 前8條

1 冉立偉;王昊;蘭東;賈紅俠;于思思;;Effects of RNA Interference Combined with Ultrasonic Irradiation and SonoV ue Microbubbles on Expression of STAT3 Gene in Keratinocytes of Psoriatic Lesions[J];Journal of Huazhong University of Science and Technology(Medical Sciences);2017年02期

2 賈曉鵬;齊春俠;路玲玲;魏民;;卷曲螺旋結(jié)構(gòu)蛋白8(CCDC8)功能研究進(jìn)展[J];生命的化學(xué);2016年05期

3 Anja Geisler;Henry Fechner;;MicroRNA-regulated viral vectors for gene therapy[J];World Journal of Experimental Medicine;2016年02期

4 孟凡榮;陳琛;萬海粟;周清華;;慢病毒載體及其研究進(jìn)展[J];中國肺癌雜志;2014年12期

5 張婷;陶春英;李云芬;楊曉娟;聶建云;;CCDC8基因的研究現(xiàn)狀[J];中國醫(yī)藥指南;2013年19期

6 李秀梅;尤玉琴;劉光澤;黎經(jīng)緯;陳媚娟;陳文吟;周軍輝;孔祥平;;慢病毒介導(dǎo)的綠色熒光蛋白轉(zhuǎn)基因小鼠的制備及鑒定[J];解放軍醫(yī)學(xué)雜志;2011年09期

7 ;Lentivirus-mediated RNA Interference and Over-expression of CDK2AP1 CDNA Regulate CDK2AP1 Expression in Human Lung Cancer A549 Cells[J];Chemical Research in Chinese Universities;2011年03期

8 ;Lentivirus-mediated shRNA interference targeting STAT3 inhibits human pancreatic cancer cell invasion[J];World Journal of Gastroenterology;2009年30期

相關(guān)碩士學(xué)位論文 前1條

1 張婷;CCDC8與ANKRA2基因在乳腺癌組織中的表達(dá)及其臨床意義[D];昆明醫(yī)科大學(xué);2013年

，

本文編號(hào)：1748052