Construction and heterologous expression of a truncated Haemagglutinin (HA) protein from the avian influenza virus H5N1 in Escherichia coli
- Author:
Chee Wei, T.
;
Nurul Wahida, A.G.
;
Shaharum, S.
- Publication Type:Journal Article
- From:Tropical Biomedicine
2014;31(4):792-801
- CountryMalaysia
- Language:English
-
Abstract:
Malaysia first reported H5N1 poultry case in 2004 and subsequently outbreak in
poultry population in 2007. Here, a recombinant gene encoding of peptide epitopes, consisting
fragments of HA1, HA2 and a polybasic cleavage site of H5N1 strain Malaysia, was amplified
and cloned into pET-47b(+) bacterial expression vector. DNA sequencing and alignment
analysis confirmed that the gene had no alteration and in-frame to the vector. Then, Histagged
truncated HA protein was expressed in Escherichia coli BL21 (DE3) under 1 mM IPTG
induction. The protein expression was optimized under a time-course induction study and
further purified using Ni-NTA agarose under reducing condition. Migration size of protein was
detected at 15 kDa by Western blot using anti-His tag monoclonal antibody and demonstrated
no discrepancy compared to its calculated molecular weight.
- Full text:W020150602331556249144.pdf
