TALEN介导的CXCR4敲除肝癌细胞株的建立

doi:10.13560/j.cnki.biotech.bull.1985.2016.02.031

生物技术通报 ›› 2016, Vol. 32 ›› Issue (2): 225-228.doi: 10.13560/j.cnki.biotech.bull.1985.2016.02.031

TALEN介导的CXCR4敲除肝癌细胞株的建立

张文美^1，2, 丁妍¹, 郭兴荣¹, 李东升¹, 赵万红², 王小莉¹

1.湖北医药学院附属太和医院胚胎干细胞研究湖北省重点实验室，十堰 442000；2.湖北医药学院生物医学工程学院，十堰 442000

收稿日期:2015-05-12 出版日期:2016-02-24 发布日期:2016-02-25
作者简介:张文美，女，研究方向：肿瘤的基因治疗；E-mail：1436271046@qq.com
基金资助:
湖北医药学院自然科学研究启动金(2011QDZR-26)，国家级大学生创新创业训练计划项目(201410929004)

The Establishment of HepG2 Cell Line with TALEN-mediated Knockout of CXCR4

ZHANG Wen-mei^1，2, DING Yan¹, GUO Xing-rong¹, LI Dong-sheng¹, ZHAO Wan-hong², WANG Xiao-li¹

(1.Hubei Key Laboratory of Embryonic Stem Cell Research，Hubei University of Medicine，Shiyan 442000；2.College of Biomedical Engineering，Hubei University of Medicine，Shiyan 442000)

Received:2015-05-12 Published:2016-02-24 Online:2016-02-25

摘要/Abstract

摘要： 通过TALEN打靶建立CXCR4的细胞株，旨在研究CXCR4对肝癌的影响。选用肝癌细胞株HepG2，采用转录激活样效应物核酸酶(TALEN)干扰细胞中CXCR4的表达。构建的CXCR4 TALEN质粒转入HepG2细胞，通过T7E1酶切确定打靶效率为40%，并通过测序筛选出了CXCR4敲除的单克隆细胞，免疫荧光和Western blot进一步证实CXCR4基因表达显著下调。

关键词: TALEN, CXCR4, 肝癌, 基因打靶

Abstract: It was to research on the effect of CXCR4 on HepG2 cells, we established one cell line which could down-regulate CXCR4 expression stably by transcription activaor-like effector nuclease (TALEN) gene targeting.The hepatocellular carcinoma cell line HepG2 was chosen for this study, and the expression of CXCR4 was interfered by transcription activator-like effector nuclease (TALEN).The constructed CXCR4 TALEN plasmids were transfected into HepG2 cells, and the targeting efficiency was determined as 40% by enzyme digestion of T7E1.The single clone cell of CXCR4 knockout was screened by gene sequencing, and the significant decrease of gene CXCR4 expression was confirmed by immunofluorescence and Western blot.The hepatocellular carcinoma cell line with stably down-regulating CXCR4 expression was established successfully by TALEN gene targeting.

Key words: TALEN, CXCR4, hepatocellular carcinoma cell, gene target

张文美, 丁妍, 郭兴荣, 李东升, 赵万红, 王小莉. TALEN介导的CXCR4敲除肝癌细胞株的建立[J]. 生物技术通报, 2016, 32(2): 225-228.

ZHANG Wen-mei, DING Yan, GUO Xing-rong, LI Dong-sheng, ZHAO Wan-hong, WANG Xiao-li. The Establishment of HepG2 Cell Line with TALEN-mediated Knockout of CXCR4[J]. Biotechnology Bulletin, 2016, 32(2): 225-228.

参考文献

[1]Tan XY, Chang S, Liu W, et al.Silencing of CXCR4 inhibits tumor cell proliferation and neural invasion in human hilar cholangiocarcinoma[J].Gut Liver, 2014, 8(2)：196-204.
[2] Hattermann K, Holzenburg E, Hans F, et al.Effects of the chemokine CXCL12 and combined internalization of its receptors CXCR4 and CXCR7 in human MCF-7 breast cancer cell[J].Cell Tissue Res, 2014, 357(1)：253-266.
[3] Ghanem I, Riveiro ME, Paradis V, et al.Insights on the CXCL12-CXCR4 axis in hepatocellular carcinoma carcinogenesis[J].Am J Transl Res, 2014, 6(4)：340-352.
[4] Sun N, Zhao H.Transcription activator-like effector nucleases(TALENs)：a highly efficient and versatile tool for genome editing[J].Biotechnol Bioeng, 2013, 110(7)：1811-1821.
[5] Cermak T, Doyle EL, Christian M, et al.Efficient design and assembly of custom TALEN and other TAL effector-based constructs for DNA targeting[J].Nucleic Acids Res, 2011, 39(12)：e82.
[6] Shinoda M, Kishida N, Itano O, et al.Long-term complete response of advanced hepatocellular carcinoma treated with multidisciplinary therapy including reduced dose of sorafenib：case report and review of the literature[J].World J Surg Oncol, 2015, 13：144.
[7] Zlotnik, A.Chemokines and cancer[J].Int J Cancer, 2006, 119(9)：2026-2029.
[8] Homey B, Müller A, Zlotnik A.Chemokines：agents for the immunotherapy of cancer?[J].Nat Rev Immunol, 2002, 2(3)：175-184.
[9] Kim J, Takeuchi H, Lam ST, et al.Chemokine receptor CXCR4 expression in colorectal cancer patients increases the risk for recurrence and for poor survival[J].J Clin Oncol, 2005, 23(12)：2744-2753.
[10] Gao Y, Li C, Nie M, et al.CXCR4 as a novel predictive biomarker for metastasis and poor prognosis in colorectal cancer[J].Tumour Biol, 2014, 35(5)：4171-4175.
[11] Zhu Y, Yang P, Wang Q, et al.The effect of CXCR4 silencing on epithelial-mesenchymal transition related genes in glioma U87 cells[J].Anat Rec(Hoboken), 2013, 296(12)：1850-1856.
[12] Liang Z, Bian X, Shim H.Inhibition of breast cancer metastasis with microRNA-302a by downregulation of CXCR4 expression[J].Breast Cancer Res Treat, 2014, 146(3)：535-542.
[13] Hiller DJ, Li BD, Chu QD.CXCR4 as a predictive marker for locally advanced breast cancer post-neoadjuvant therapy[J].J Surg Res, 2011, 166(1)：14-18.
[14] Xie L, Wei J, Qian X, et al.CXCR4, a potential predictive marker for docetaxel sensitivity in gastric cancer[J].Anticancer Res, 2010, 30(6)：2209-2216.
[15] Paratore S, Banna GL, D’Arrigo M, et al.CXCR4 and CXCL12 immunoreactivities differentiate primary non-small-cell lung cancer with or without brain metastases[J].Cancer Biomark, 2011, 10(2)：79-89.
[16] Paratore S, Banna GL, D’Arrigo M, et al.CXCR4 is a prognostic marker in acute myelogenous leukemia[J].Blood, 2007, 109(2)：786-791.
[17] Xiang ZL, Zeng ZC, Tang ZY, et al.Chemokine receptor CXCR4 expression in hepatocellular carcinoma patients increases the risk of bone metastases and poor survival[J].BMC Cancer, 2009, 9：176.
[18] Wang L, Huang T, Chen W, et al.Silencing of CXCR4 by RNA interference inhibits cell growth and metastasis in human renal cancer cells[J].Oncol Rep, 2012, 28(6)：2043-2048.

TALEN介导的CXCR4敲除肝癌细胞株的建立

The Establishment of HepG2 Cell Line with TALEN-mediated Knockout of CXCR4

PDF (PC)

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 15

编辑推荐

Metrics

本文评价

[1]	常允建, 康冉, 薛璇, 王韶畅, 赵庆文, 郭志云. 肝癌中增强子调控miRNA前馈环路的识别与功能分析[J]. 生物技术通报, 2019, 35(5): 140-145.
[2]	霍桂桃, 杨艳伟, 吴曦, 刘甦苏, 李芊芊, 周舒雅, 柳全明, 王三龙, 沈月雷, 吕建军, 范昌发. p53基因敲除大鼠模型的构建及表型分析[J]. 生物技术通报, 2018, 34(8): 170-174.
[3]	汤波,孙照霖,戴蕴平. 牛的基因编辑技术研究进展[J]. 生物技术通报, 2018, 34(5): 41-47.
[4]	毕延震,肖红卫,张立苹,任红艳,华再东,华文君,王正,牛敏杰,林郑云,任习东,孙理华,郑新民. 非人灵长类动物基因编辑技术的应用及挑战[J]. 生物技术通报, 2018, 34(5): 48-56.
[5]	张晶晶, 金小宝, 李小波, 汪洁, 马艳. 过表达Glypican-3对高转移性肝癌细胞HCCLM3生物学行为的影响[J]. 生物技术通报, 2017, 33(12): 176-184.
[6]	姚杨,苏杰,刘凯歌,徐锐. 上调基因-11(URG11)生物信息学与功能分析[J]. 生物技术通报, 2016, 32(3): 203-208.
[7]	王小莉,余庆,袁雅红，腾智平,李东升,曾毅. 利用TALEN技术建立恒河猴TRIM5α基因突变细胞株[J]. 生物技术通报, 2016, 32(12): 53-57.
[8]	李铎,李佳文,袁群琛,林川川,宋关斌. 溶血磷脂酸对不同转移性肝癌细胞迁移行为的影响[J]. 生物技术通报, 2016, 32(12): 189-194.
[9]	李斌, 许晓亚, 杨刚刚, 崔晴晴, 杨亚娟, 徐存拴. 重组人白细胞介素-1α在毕赤酵母中的表达、纯化及生物学活性检测[J]. 生物技术通报, 2015, 31(9): 244-250.
[10]	吴璐,王磊,任远,原辉. 基因组编辑技术研究进展[J]. 生物技术通报, 2014, 0(11): 84-90.
[11]	周晓明;傅海媛;黄亚娟;侯利平;孙云波;原剑;杨保安;郑俊杰;甄蓓;肖汉族;貌盼勇;魏开华;. 多肽抗体检测原发性肝癌血清标志物ELISA方法的建立及应用[J]. , 2012, 0(05): 179-184.
[12]	姚杨;苏杰;刘凯歌;徐锐;成碧萍;. 乙肝相关性肝癌中新基因CHCHD2的生物信息学分析[J]. , 2012, 0(03): 179-185.
[13]	冀颖;李梅;蒋细良;. 基因打靶与RNA干扰技术在丝状真菌基因功能组学研究中的应用[J]. , 2011, 0(05): 64-70.
[14]	王凌云;成功;周欢敏;. 动物转基因研究中的技术方法[J]. , 2010, 0(12): 73-77.
[15]	张丽娜;刘国安;杨红;. 基因打靶技术的研究进展[J]. , 2010, 0(09): 51-55.