|本期目录/Table of Contents|

[1]董 旭,张兆域,苟芳琳,等.利用CRISPR/Cas9 系统构建稳定敲除CBX2基因的A549细胞系[J].天津医科大学学报,2018,24(05):395-398.
　DONG Xu,ZHANG Zhao-yu,GOU Fang-lin,et al.Construction of CBX2 gene knockout stable A549 cell line with CRISPR /Cas9 system[J].Journal of Tianjin Medical University,2018,24(05):395-398.
点击复制

利用CRISPR/Cas9 系统构建稳定敲除CBX2基因的A549细胞系(PDF)
分享到：

《天津医科大学学报》[ISSN:1006-8147/CN:12-1259/R]

卷:
24卷
期数:
2018年05期
页码:
395-398
栏目:
基础医学
出版日期:
2018-09-20

文章信息/Info

Title:
Construction of CBX2 gene knockout stable A549 cell line with CRISPR /Cas9 system
作者:
董 旭; 张兆域; 苟芳琳; 兰 蓓
天津医科大学生物化学与分子生物学系，天津 300070
Author(s):
DONG Xu;  ZHANG Zhao-yu;  GOU Fang-lin;  LAN Bei
Department of Biochemistry and Molecular Biology, Tianjin Medical University, Tianjin 300070, China
关键词:
CBX2基因; CRISPR/Cas9 系统; 基因敲除
Keywords:
CBX2 gene;  CRISPR/Cas9 system;  gene knockout
分类号:
Q7
DOI:
-
文献标志码:
A
摘要:
目的：利用CRISPR/Cas9系统构建稳定敲除CBX2基因的A549细胞系。方法：设计针对敲除CBX2的short guide RNA（sgRNA），并克隆到载体PX459中。将测序正确的重组质粒转染到A549细胞中，利用嘌呤霉素筛选转染阳性细胞并分离得到5个单克隆细胞系，通过Western blot方法检测构建的细胞系中CBX2蛋白表达情况。结果：成功构建了靶向CBX2的CRISPR/Cas9重组质粒，筛选出的5个单克隆细胞系中CBX2蛋白表达水平均显著下降。结论：成功利用CRISPR/Cas9 系统构建了稳定敲除CBX2基因的A549细胞系。
Abstract:
Objective: To construct CBX2 gene knockout stable A549 cell line with CRISPR/Cas9 system. Methods：The single guide RNA (sgRNA) targeting CBX2 gene was designed and inserted into the expression vector PX459 using molecular cloning technique. The recombined plasmid was verified by sequencing. The plasmid was transfected into A549 cell line. After puromycine selection, monoclonal cell line was picked for expansion. Finally, the expression of CBX2 protein in A549 CBX2 knockout cell line was detected by Western blot. Results：The CRISPR / Cas9 recombinant plasmid targeting CBX2 was successfully constructed, and the expression of CBX2 protein was significantly decreased. Conclusion：The A549 CBX2 knockout cell line could be successfully constructed by CRISPR/Cas9 system.

参考文献/References:


[1] Morey L，Helin K. Polycomb group protein-mediated repression of transcription[J]. Trends Biochem Sci，2010，35（6）：323
[2] Muller J，Verrijzer P. Biochemical mechanisms of gene regulation by polycomb group protein complexes[J]. Curr Opin Genet Dev，2009， 19（2）：150
[3] Pirrotta V. Polycombing the genome： PcG，trxG，and chromatin silencing[J]. Cell，1998，93（3）：333
[4] Sauvageau M，Sauvageau G. Polycomb group proteins： multi-faceted regulators of somatic stem cells and cancer[J]. Cell Stem Cell，2010， 7（3）：299
[5] Pirrotta V，Poux S，Melfi R，et al. Assembly of Polycomb complexes and silencing mechanisms[J]. Genetica，2003，117（2/3）：191
[6] Bezsonova I，Walker J R，Bacik J P，et al. Ring1B contains a ubiquitin-like docking module for interaction with Cbx proteins[J]. Biochemistry，2009，48（44）：10542
[7] Li J，Xu Y，Long X D，et al. Cbx4 governs HIF-1alpha to potentiate angiogenesis of hepatocellular carcinoma by its SUMO E3 ligase activity[J]. Cancer Cell，2014，25（1）：118
[8] Wang X，Li L，Wu Y，et al. CBX4 Suppresses Metastasis via Recruitment of HDAC3 to the Runx2 Promoter in Colorectal Carcinoma[J]. Cancer Res，2016，76（24）：7277
[9] Santanach A，Blanco E，Jiang H，et al. The Polycomb group protein CBX6 is an essential regulator of embryonic stem cell identity[J]. Nat Commun，2017，8（1）：1235
[10] Forzati F，Federico A，Pallante P，et al. CBX7 is a tumor suppressor in mice and humans[J]. J Clin Invest，2012，122（2）：612
[11] Forzati F，Federico A，Pallante P，et al. Tumor suppressor activity of CBX7 in lung carcinogenesis[J]. Cell Cycle，2012，11（10）：1888
[12] Tan J，Jones M，Koseki H，et al. CBX8，a polycomb group protein，is essential for MLL-AF9-induced leukemogenesis[J]. Cancer Cell，2011，20（5）：563
[13] Sapranauskas R，Gasiunas G，Fremaux C，et al. The Streptococcus thermophilus CRISPR/Cas system provides immunity in Escherichia coli[J]. Nucleic Acids Res，2011，39（21）：9275
[14] Terns M P，Terns R M. CRISPR-based adaptive immune systems[J]. Curr Opin Microbiol，2011，14（3）：321
[15] Cong L，Ran F A，Cox D，et al. Multiplex genome engineering using CRISPR/Cas systems[J]. Science，2013，339（6121）：819
[16] Core N，Joly F，Boned A，et al. Disruption of E2F signaling suppresses the INK4a-induced proliferative defect in M33-deficient mice[J]. Oncogene，2004，23（46）：7660
[17] van den Boom V，Rozenveld-Geugien M，Bonardi F，et al. Nonredundant and locus-specific gene repression functions of PRC1 paralog family members in human hematopoietic stem/progenitor cells[J]. Blood，2013，121（13）：2452

相似文献/References:

备注/Memo

备注/Memo:
-
更新日期/Last Update: 2018-09-30