|
|
Expression,Purification and Antibody Preparation of Recombinant C8orf32 Protein |
|
|
Abstract C8orf32 is a gene which has not been functionally characterized,the mRNA level of this gene is significantly higher in breast cancer tissues than that in normal breast tissues.The amplified cDNA fragment was inserted into the pGEX6P1 vector fused with the upstream GST gene.The expression vector was transformed into the E.coli BL21(DE3) strain and expression of GST C8orf32 fusion protein was induced by IPTG..After removal of GST tag by sitespecific protease,the C8orf32 protein fused with an eight amino acid peptide tag was obtained.The purity of recombinant C8orf32 protein was about 95%.The identity of the purified protein was confirmed by Nterminal sequencing and tandem mass spectrometry.The polyclonal antibody was prepared by immunizing the New Zealand white rabbits with C8orf32 protein.The polyclonal antibody was proved to recognize the C8orf32 protein correctly.The purified C8orf32 protein can be used for structural and functional studies and the polyclonal antibody can be used for tissue specific protein expression profiling.
|
Received: 11 November 2008
Published: 27 April 2009
|
|
|
|
[1] Ota T,Suzuki Y.Complete sequencing and characterization of 21,243 fulllength human cDNAs.Nat Genet,2004,36(1): 40~45
[2] Tian L,Wang P,Shi T,et al.Screening for novel human genes associated with CRE pathway activation with cell microarray.Genomics,2007,90(1): 28~34
[3] Lim J,Hao T,Shaw C,et al.A proteinprotein interaction network for human inherited ataxias and disorders of Purkinje cell degeneration.Cell,2006,125(4): 801~814
[4] Smith D B,Johnson K S.Singlestep purification of polypeptides expressed in Escherichia coli as fusions with glutathione Stransferase.Gene,1988,67(1): 31~40
[5] Frangioni J V,Neel B G.Solubilization and purification of enzymatically active glutathione Stransferase (pGEX) fusion proteins.Anal.Biochem,1993,210(1): 179~187
[6] 李赛男,李朝飞,潘丽晶,等.斜纹夜蛾核多角体病毒VP39 GST融合蛋白原核表达.中国生物工程杂志,2006,26(9):16~19
Li S N,Li Z F,Pan L J,et al.China Biotechnology,2006,26(9):16~19
[7] 蔡仕英,马贤凯.提纯性融合蛋白.中国生物工程杂志,1988,1(26):41~48
Cai S Y,Ma X K.China Biotechnology,1988,1(26):41~48
[8] 李伟.重组人MBD4蛋白在大肠杆菌中的表达、纯化及活性分析.中国生物化学与分子生物学报,2006,22(12): 979~983
Li W.Chinese Journal of Biochemistry and Molecular Biology,2006,22(12): 979~983
|
|
Viewed |
|
|
|
Full text
|
|
|
|
|
Abstract
|
|
|
|
|
Cited |
|
|
|
|
|
Shared |
|
|
|
|
|
Discussed |
|
|
|
|