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Prokaryotic Expression System Construction of the Tufted Deer(Elaphodus cephalophus)AIF-1(PDF)


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Prokaryotic Expression System Construction of the Tufted Deer(Elaphodus cephalophus)AIF-1
Zhou NannanYang ZhenLuo LiyunSong FeiBai MinCao Xiangrong
School of Life Sciences,Nanjing Normal University,Jiangsu Key Laboratory for Molecular and Medical Biotechnology,Nanjing 210023,China
Tufted deer(Elaphodus cephalophus)AIF-1prokaryotic expression
The TdAIF鄄1 cDNA was cloned from the testis cDNA library of the Tufted deer(Elaphodus cephalophus) and analyzed by bioinformatic methods. Primers were designed according to cDNA sequence to clone the gene. The gene was cloned into pMD19鄄T vector for sequencing. The right sequence was digested by restriction enzyme and subcloned into the expression vector pET鄄28a( +). After transformed into E. coli BL21( DE3),the recombinant plasimid was induced to express by IPTG. The E. coli BL21(DE3)expressed recombinant protein was broken by ultrasonic to show whether the re鄄 combinant protein was soluble or not. Lastly,the soluble protein was purified. The recombinant protein was analyzed and identificated by SDS-PAGE electrophoresis and Western blot. Analysis of sequence showed that the TdAIF-1 cDNA contained a 438 bp op-n reading frame encoding 145 amino acids. The recombinant plasimid was correctly constructed according to sequencing and restriction enzyme analysis. The recombinant protein was about 20 kD and soluble mainly. When the recombinant protein was purified,using elution buffer containing 50 mmol/ L or 100 mmol/ L imidazole could get purified protein. The TdAIF-1 Prokaryotic Expression System was constructed successfully and recombinant protein was obtained,which was helpful for the future study of its biological function.


[1]Utans U,Arceci R J,Yamashita Y,et al. Cloning and characterization of allograft inflammatory factor-1:a novel macrophage factor identified in rat cardiac allografts with chronic rejection[ J]. Journal of Clinical Investigation,1995,95(6):2 954-2 962.
[2] Imai Y,Kohsaka S. Intracellular signaling in M-CSF-induced microglia activation:role of Iba1[ J]. Glia,2002,40(2):164-174.
[3] Kruse M,Steffen R,Batel R,et al. Differential expression of allograft inflammatory factor 1 and of glutathione peroxidase during auto-and allograft response in marine sponges[J]. Journal of Cell Science,1999,112(23):4 305-4 313.
[4] De Zoysa M,Nikapitiya C,Kim Y,et al. Allograft inflammatory factor-1 in disk abalone( Haliotis discus discus):Molecular cloning,transcriptional regulation against immune challenge and tissue injury[ J]. Fish and Shellfish Immunology,2010,29(2):319-326.
[5] Fujiki K,Shin D H,Nakao M,et al. Molecular cloning of carp(Cyprinus carpio) CC chemokine,CXC chemokine receptors,allograft inflammatory factor-1,and natural killer cell enhancing factor by use of suppression subtractive hybridization[J]. Im-munogenetics,1999,49:909-914.
[6] Watano K,Iwabuchi K,Fujii S,et al. Allograft inflammatory factor-1 augments production of interleukin-6,-10 and -12 by a mouse macrophage line[J]. Immunology,2001,104(3):307-316.
[7] Utans U, Quist W C, McManus B M, et al. Allograft inflammatory factory-1: a cytokine-responsive macrophage molecule expressed in transplanted human hearts[J]. Transplantation,1996,61(9):1 387-1 392.
[8]Ohsawa K,Imai Y,Kanazawa H,et al. Involvement of Iba1 in membrane ruffling and phagocytosis of macrophages/ microglia[J]. Journal of Cell Science,2000,113(17):3 073-3 084.
[9] Sasaki Y,Ohsawa K,Kanazawa H,et al. Iba1 is an actin-cross-linking protein in macrophages/ microglia[J]. Biochemical and Biophysical Research Communications,2001,286:292-297.
[10] Schulze J O,Quedenau C,Roske Y,et al. Structural and functional characterization of human Iba proteins[J]. FEBS Journal, 2008,275(18):4 627-4 640.
[11] Iida H,Doiguchi M,Yamashita H,et al. Spermatid-specific expression of Iba1,an ionized calcium binding adapter molecule-1,in rat testis[J]. Biology of Reproduction,2001,64(4):1 138-1 146.
[12] Autieri M V,Carbone C,Mu A. Expression of allograft inflammatory factor-1 is a marker of activated human vascular smooth muscle cells and arterial injury[J]. Arteriosclerosis,Thrombosis,and Vascular Biology,2000,20:1 737-1 744.
[13] Autieri M V,Kelemen S E,Wendt K W. AIF-1 is an actin-polymerizing and Rac1-activating protein that promotes vascular smooth muscle cell migration[J]. Circulation Research,2003,92(10):1 107-1 114.
[14] Chen X,Kelemen S E,Autieri M V. AIF-1 expression modulates proliferation of human vascular smooth muscle cells by autocrine expression of G-CSF[J]. Arteriosclerosis,Thrombosis,and Vascular Biology,2004,24:1 217-1 222.
[15] 曹祥荣,束峰钰,张锡然,等. 毛冠鹿与3 种麂属动物的线粒体细胞色素b 的系统进化分析[ J]. 动物学报,2002,48(1):44-49.
[16] 张文,汤文文,庞宏,等. 毛冠鹿睾丸组织cDNA 文库的构建及P1 精蛋白基因的克隆[J]. 兽类学报,2006,26(2):164-170.
[17] Tian Y,Autieri M V. Cytokine expression and AIF-1-mediated activation of Rac2 in vascular smooth muscle cells:a role for Rac2 in VSMC activation[J]. American Journal of Physiology-Cell Physiology,2007,292(2):841-849.


Last Update: 2014-06-30