Journal of Veterinary Research

Journal of Veterinary Research

CLONING, SEQUENCINGAND EXPRESSION OF HSP70 C- TERMINALDOMAIN IN MYCOBACTERIUM AVIUM SUBSP. PARATUBERCULOSIS

Authors
gholamraza nikbakhat
Gholam Reza Nikbakht Broujeni
Hasan Tadjbakhsh
Nader Mosavari
Syed Amir Hossein Jalali
Abstract
Heat shock proteins (HSPs) have been shown to act as an adjuvant when co-administered with
different antigens, especially tumor antigens or antigens from infectious agents. C-terminal
domain of Mycobacterium tuberculosis heat shock protein 70 (Hsp70), when fused to peptide
antigens, provides a unique structure that is able to induce potent immune responses. In this study,
aneukaryotic expression vector pEGFP-N1, containing C-terminal domain of Mycobacterium
paratuberculosis HSP 70, Green Fluorescent Protein (GFP) gene in the plasmid construct , was
designed for use as a reporter. With GFP system, expression of the target protein was evaluated
in the cell culture. The nucleotide sequence of the cloned gene was revealed by sequencing. The
protein expression of designed plasmid was also proved by reverse transcriptase polymerase
chain reaction (RT-PCR). Our eukaryotic expression vector (pEGFP-N1 -hsp70 c-terminal) was
successfully constructed and HSP70 c-terminal domain protein was expressed effectively. The
current experiment, as a basis for a new DNAvaccine design, can be used for the future studies on
reverse vaccinology
Keywords
green fluorescent protein (GFP).
heat shock protein 70(Hsp70)
Mycobacterium avium subsp. paratuberculosis
Journal of Veterinary Research
Volume 66, Issue 4 - Serial Number 4
Autumn 2011
Pages 289-298