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Secreted expression of the classical swine fever virus glycoprotein E(rns) in yeast and application to a sandwich blocking ELISA.

Abstract：

:E(rns) is an envelope glycoprotein of classical swine fever virus (CSFV) with RNase activity. The purpose of this study was to produce an active E(rns) for further applications using the yeast secreted expression system. The E(rns) gene was cloned into the expression vector pGAPZalphaC which was introduced into Pichia pastoris. Expression of E(rns) protein in culture supernatant was confirmed by Western blot analysis using both the monoclonal antibody against CSFV E(rns) and CSFV-positive swine serum. The yeast-expressed E(rns) (yE(rns)) was shown to have N-linked glycosylation and to form homodimer of 74 kDa molecules. All monomer, homodimer, and deglycosylated forms of yE(rns) demonstrated intrinsic ribonuclease activity and a clear preference for uridine-rich sequence. A direct sandwich blocking enzyme-linked immunosorbent assay (ELISA) based on the yE(rns) was developed with a high sensitivity and specificity. The yE(rns) which possesses enzymatic activity and retains antigenicity may provide a useful material for developing a diagnostic kit.

journal_name

J Virol Methods

journal_title

Journal of virological methods

authors

Huang C,Chien MS,Hu CM,Chen CW,Hsieh PC

doi

10.1016/j.jviromet.2005.08.020

subject

Has Abstract

pub_date

2006-03-01 00:00:00

pages

40-7

issue

1-2

eissn

0166-0934

issn

1879-0984

pii

S0166-0934(05)00285-5

journal_volume

132

pub_type

杂志文章

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