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2013•Nanjing Nongye Daxue xuebaoRequires access

Construction and identification of phage ScFv library against fumonisin B_1

Yin Wang

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Abstract

The VH and VL gene fragments were first amplified from hybridoma cell line(F3) specific to fumonisin B1(FB1) integrated as single-chain antibody(ScFv) gene using gene splicing by overlap extension PCR(SOE-PCR).The ScFv was cloned into a phagemid vector pCANTAB 5E,and then was transformed into Escherichia coli TG1 to establish a phage ScFv library with the helper phage M13K07.The results indicated that the insert rate of SvFv gene was 100%,the phage antibody contained 1.5×107independent clones and phage titer was 2.1×1015PFU·mL-1.After biopanning enrichment and screening,five strains that could secrete the FB1 specific phage single-chain antibody were selected successfully.Conclusion: the phage ScFv library against FB1 was constructed successfully.

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What this paper is about

The VH and VL gene fragments were first amplified from hybridoma cell line(F3) specific to fumonisin B1(FB1) integrated as single-chain antibody(ScFv) gene using gene splicing by overlap extension PCR(SOE-PCR).The ScFv was cloned into a phagemid vector pCANTAB 5E,and then was transformed into Escherichia coli TG1 to establish a phage ScFv library with the helper phage M13K07.The results indicated that the insert rate of SvFv gene was 100%,the phage antibody contained 1.5×107independent clones and phage titer was 2.1×1015PFU·mL-1.After biopanning enrichment and screening,five strains that could secrete the FB1 specific phage single-chain antibody were selected successfully.Conclusion: the phage ScFv library against FB1 was constructed successfully.

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Available abstract

The VH and VL gene fragments were first amplified from hybridoma cell line(F3) specific to fumonisin B1(FB1) integrated as single-chain antibody(ScFv) gene using gene splicing by overlap extension PCR(SOE-PCR).The ScFv was cloned into a phagemid vector pCANTAB 5E,and then was transformed into Escherichia coli TG1 to establish a phage ScFv library with the helper phage M13K07.The results indicated that the insert rate of SvFv gene was 100%,the phage antibody contained 1.5×107independent clones and phage titer was 2.1×1015PFU·mL-1.After biopanning enrichment and screening,five strains that could secrete the FB1 specific phage single-chain antibody were selected successfully.Conclusion: the phage ScFv library against FB1 was constructed successfully.

Key concepts: Biopanning, Phagemid, Phage display, Molecular biology, Biology, Gene, Insert (composites), Antibody

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