title: [Construction and screening of SARS-CoV S protein-specific phage displayed antigen library].
abstract: The aim of this study is to construct a SARS-CoV S protein-specific phage displayed antigen library for the epitope characterization of anti-S monoclonal antibodies (mAbs). First, the full-length gene of SARS-S protein was PCR amplified, purified and then digested with DNase I to obtain DNA fragments in the size range of 50-500 bp. The resulting fragments were blunt-end ligated to the modified phage display vector pComb3XSS. The reactions were electrotransformed into XL1-Blue and infected with VCSM13 helper phage. The SARS-CoV S protein-specific phage displayed antigen library was biopanned and screened against two anti-S mAbs, S-M1 and S-M2. The results showed that we successfully constructed the phage displayed antigen library with a size of 5.7 x 10(6). After three-rounds of biopanning, 14 positive phage clones for S-M1 and 15 for S-M2 were respectively identified. Sequence analyses revealed the possible epitopes of two mAbs. Therefore, the S protein-specific phage displayed antigen library provides a crucial platform for the epitope characterization of anti-S antibodies and it is highly valuable for development of SARS vaccines and diagnostics.
authors.affiliation: Institute of Medical Virology, School of Laboratory Medicine, Wenzhou Medical College, Wenzhou 325035, China. wuruiping_q@126.com
authors.forename: Rui-Ping
authors.initials: RP
authors.lastname: Wu
authors.forename: Jia-Zi
authors.initials: JZ
authors.lastname: Meng
authors.forename: Yu-Xian
authors.initials: YX
authors.lastname: He
mesh_terms: D000914:Antibodies, Viral / Q000276:immunology; D001435:Bacteriophages / Q000235:genetics* / Q000378:metabolism; D000939:Epitopes / Q000235:genetics / Q000276:immunology; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics* / Q000276:immunology; D019151:Peptide Library; D045473:Severe acute respiratory syndrome-related coronavirus / Q000235:genetics* / Q000276:immunology; D045169:Severe Acute Respiratory Syndrome / Q000276:immunology / Q000821:virology*; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000276:immunology
chemical_list: D000914:Antibodies, Viral; D000939:Epitopes; D008562:Membrane Glycoproteins; D019151:Peptide Library; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins; C578557:spike glycoprotein, SARS-CoV; C578553:spike protein, mouse hepatitis virus