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Expression of bovine interferon-γ in Escherichia coli and Pichia pastoris and comparison of their antiviral activities

Published online by Cambridge University Press:  12 February 2007

Shi Xi-Ju ,
Zhang Can ,
Han Chun-Lai ,
Xia Chun  and
Wang Ming
Shi Xi-Ju*
Affiliation:
College of Veterinary Medicine, China Agricultural University, Beijing 100094, China
Zhang Can
Affiliation:
College of Veterinary Medicine, China Agricultural University, Beijing 100094, China
Han Chun-Lai
Affiliation:
College of Veterinary Medicine, China Agricultural University, Beijing 100094, China
Xia Chun
Affiliation:
College of Veterinary Medicine, China Agricultural University, Beijing 100094, China
Wang Ming*
Affiliation:
College of Veterinary Medicine, China Agricultural University, Beijing 100094, China
*
**Corresponding author. E-mail: [email protected]
*Current address: Beijing Entry-exit Inspection and Quarantine Bureau
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Abstract

The bovine interferon-γ gene cloned from total RNA of peripheral blood lymphocytes stimulated with concanavalin A (ConA), using the reverse transcriptase-polymerase chain reaction (RT-PCR), was cloned into pGEM T-easy vector and sequenced. The mature peptide without signal peptide was then subcloned and constructed into the expression plasmids pET28a/BoIFN-γ and pPICZα/BoIFN-γ, respectively. The former was highly expressed, induced by isopropyl β-d-thiogalactoside (IPTG), in Escherichia coli BL21, with 30% of bacterial proteins in inclusion bodies. The latter was expressed in Pichia pastoris GS115 with methanol induction. The expressed products were secreted directly into cultural supernatant at concentrations of 1.0 g/l. The same recombinant proteins had antiviral activities 10 times higher in an MDBK (Madan-Darby bovine kidney)/VSV (Vesicular stomatitis virus) cell line than in a CEF (chicken embryo fibroblast)/VSV) cell line, and the antiviral activities expressed in P. pastoris were higher than those expressed in E. coli in the same cell line.

Keywords

antiviral activitiesbovine IFN-γEscherichia colimolecular cloningPichia pastoris
Type
Research Article
Information
Chinese Journal of Agricultural Biotechnology , Volume 3 , Issue 1 , April 2006 , pp. 39 - 46
Copyright
Copyright © China Agricultural University and Cambridge University Press 2006

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