水牛转录抑制因子CTCF基因克隆分析及不同组织中的表达研究

›› 2013, Vol. 40 ›› Issue (3): 1-6.

• 生物技术 • 下一篇

水牛转录抑制因子CTCF基因克隆分析及不同组织中的表达研究

苏节, 朱鹏, 刘庆友, 雷小灿, 石德顺, 崔奎青

广西大学,亚热带生物资源保护利用国家重点实验室,广西南宁 530004

收稿日期:2012-07-26 出版日期:2013-03-20 发布日期:2013-03-19
通讯作者: 石德顺,研究员,博士导师。E-mail:ardsshi@gxu.edu.c;崔奎青,副教授,硕士导师。E-mail:kqcui@126.com E-mail:ardsshi@gxu.edu.c;kqcui@126.com
作者简介:苏节(1987-),男,广西人,硕士生,研究方向:动物生物技术。
基金资助:
国家转基因重大专项(2011ZX08007-003);国家高技术研究发展计划(863)项目(2011AA100607)。

Cloning of Buffalo CTCF Gene and its Expression Pattern Investigation in Different Tissues

SU Jie, ZHU Peng, LIU Qing-you, LEI Xiao-can, SHI De-shun, CUI Kui-qing

State Key Laboratory of Conservation and Utilization of Subtropical Agro-bioresources, Guangxi University, Nanning 530004, China

Received:2012-07-26 Online:2013-03-20 Published:2013-03-19

摘要/Abstract

摘要： 本研究克隆了水牛转录抑制因子CTCF基因序列,并运用生物信息学方法对其核苷酸序列的保守性和氨基酸的理化性质、蛋白质结构进行了系统分析,此外还对CTCF基因在水牛不同组织中的表达差异进行了检测。结果表明,应用RT-PCR技术克隆获得了长2239 bp水牛CTCF基因序列,其中编码区全长2184 bp,编码727个氨基酸,理论蛋白质分子质量82.7 ku,等电点为6.57。多重序列比较分析显示,水牛CTCF核苷酸序列与牛、猪、马、人和小鼠相应序列的相似性分别为99%、96%、96%、94%和92%,结合系统进化树分析结果推测,CTCF基因在不同物种及进化的过程中具有高度的保守性。对水牛CTCF蛋白的二级和三级结构分析结果发现,其存在连续11个锌指C2H2结构,预测其为重要的DNA结合蛋白。定量表达分析结果显示,CTCF在水牛肝脏组织中相对表达量最高,大脑、肌肉和肾脏次之,卵巢和皮肤表达量较低。

关键词: 水牛; CTCF基因; 克隆; 表达

Abstract: Buffalo CTCF gene was cloned in the present study, the CTCF sequence was systemically analysised by bioinformatics techniques, and the expression patterns of CTCF in different tissues were also assayed with Real-time fluorescence quantitative-PCR(QRT-PCR). The results showed that, with RT-PCR a 2239 bp buffalo CTCF gene fragment was cloned and sequenced, including the whole CDS of 2184 bp (coded 727 amino acids). The molecular weight and isoelectric point of buffalo CTCF protein were predicted as 82.7 ku and 6.57 respectively. The sequence multialigned results showed that buffalo CTCF gene shared 99%, 96%, 96%, 94% and 92% of similar nucleotide sequence with Bos taurus, Equus caballus, Sus scrofa, Homo sapiens and Mus musclus,respectively. It was predicted that buffalo CTCF protein contained eleven ZnF_C2H2 domains, which meaned that it was a DNA binding protein. In addition, we also analyzed the tissue expression of CTCF gene through QRT-PCR. The results showed the mRNA of CTCF gene existed in all the six detected buffalo tissues with the most abundant expression in liver, followed by brain, muscle, spleen and ovary, the minimal expression in skin was observed.

Key words: buffalo; CTCF gene; cloning; expression

中图分类号:

Q785

苏节, 朱鹏, 刘庆友, 雷小灿, 石德顺, 崔奎青. 水牛转录抑制因子CTCF基因克隆分析及不同组织中的表达研究[J]. , 2013, 40(3): 1-6.

SU Jie, ZHU Peng, LIU Qing-you, LEI Xiao-can, SHI De-shun, CUI Kui-qing. Cloning of Buffalo CTCF Gene and its Expression Pattern Investigation in Different Tissues[J]. , 2013, 40(3): 1-6.

参考文献

1 Balakrishnan S K, Witcher M, Berggren T W. Functional and molecular characterization of the role of CTCF in human embryonic stem cell biology[J]. PLoS One,2012,7(8):e42424.
2 Burke L J, Hollemann T, Pieler T. Molecular cloning and expression of the chromatin insulator protein CTCF in Xenopus laevis[J].Mech Dev,2002,113(1):95～98.
3 Chao W, Huynh K D, Spencer R J, et al. CTCF, a candidate trans-acting factor for X-inactivation choice[J]. Science,2002,295(5553):345～347.
4 Cuddapah S, Jothi R, Schones D E, et al. Global analysis of the insulator binding protein CTCF in chromatin barrier regions reveals demarcation of active and repressive domains[J].Genome Res,2009,19(1):24～32.
5 Didych D A, Kotova E S, Akopov S B, et al. DNA fragments binding CTCF in vitro and in vivo are capable of blocking enhancer activity[J]. BMC Res Notes,2012,5(5):178.
6 Filippova G N,Fagerlie S,Klenova E M,et al. An exceptionally conserved transcriptional repressor,CTCF,employs different combinations of zinc fingers to bind diverged promoter sequences of avian and mammalian c-myc oncogenes[J].Molecular and Cellular Biology,1996,16(6):2802～2813.
7 Gombert W M, Krumm A.Targeted deletion of multiple CTCF-binding elements in the human c-myc gene reveals a requirement for CTCF in C-MYC expression[J]. PLoS One,2009,4(7):e6109.
8 Klenova E M, Nicolas R H, Paterson H F, et al. CTCF,a conserved nuclear factor required for optimal transcriptional activity of the chicken c-myc gene, is an 11-Zn-finger protein differentially expressed in multiple forms[J]. Mol Cell Biol, 1993, 13(12):7612～7624.
9 Lutz M, Burke L J, Barreto G, et al. Transcriptiona repression by the insulator protein CTCF involves histone deacetylases[J]. Nucl Acids Res,2000, 28(8): 1707～1713.
10 Ohlsson R, Renkawitz R, Lobanenkov V. CTCF is a uniquely versatile transcription regulator linked to epigenetics and disease[J]. Trends Genet,2001,17(9):520～527.
11 Pabo C O, Peisach E, Grant R A. Design and selection of novel Cys2His2 zinc finger proteins[J]. Annual Review of Biochemistry,2001,70: 313～340.
12 Reik W,Murrell A. Genomic imprinting: Silence across the border[J]. Nature,2000,405(6785):408.
13 Rubio E D, Reiss D J, Welcsh P L, et al. CTCF physically links cohesion to chromatin[J]. Proc Natl Acad Sci USA,2008,105(24):8309～8314.
14 Saitoh N,Bell A C,Recillas-Targa F,et al. A region to the N-terminal side of the CTCF zinc finger domain is essential for activating transcription from the amyloid precursor protein promoter[J].The Journal of Biological Chemistry, 2002, 277: 1619～1627.
15 Uenishi H, Eguchi T, Suzuki K, et al. PEDE (pig EST data explorer): Construction of a database for ESTs derived from porcine full-length cDNA libraries[J]. Nucleic Acids Res,2004,32(Suppl.1): D484～D488.
16 Vostrov A A, Quitschke W W. The zinc finger protein CTCF binds to the APBbeta domain of the amyloid beta-protein precursor promoter. Evidence for a role in transcriptional activation[J]. J Biol Chem,1997, 272(52):33353～33359.

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水牛转录抑制因子CTCF基因克隆分析及不同组织中的表达研究

Cloning of Buffalo CTCF Gene and its Expression Pattern Investigation in Different Tissues

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