Cloning and expression of hepatitis C virus core protein in pGemex-1 expression vector.
BACKGROUND
Hepatitis C virus is one of the main causes of chronic hepatitis in developing countries. There are 170 million affected people around the world as reported by the World Health Organization. The treatment of hepatitis C is not successful in most cases; it is extremely costly, and requires prolonged therapy, therefore it is desirable to develop a vaccine to prevent the spread of hepatitis C virus.


METHODS
Hepatitis C virus RNA was extracted from a hepatitis C virus-infected serum sample. cDNA was synthesized and the hepatitis C virus core gene was amplified by polymerase chain reaction. The polymerase chain reaction product was cloned in pGEMEX-1 expression vector and expressed in Escherichia coli BL21 strain with DE3 (a lambda prophage carrying the T7 RNA polymerase gene) host by induction of promoter using one millimolar isopropyl beta-D-thiogalactopyranoside in laboratory scale. Induced lysate cells were electrophoresed on SDS- polyacrylamide gel.


RESULTS
A protein band was detected in induced cells in comparison with non-induced cells. Expressed protein was confirmed by gel diffusion and dot blot analysis using induced lysate cells as antigen and hepatitis C virus-infected serum as antibody.


CONCLUSIONS
In present study, we have provided a recombinant plasmid based on hepatitis C virus core gene.
Kazemi
Bahram
B
Cellular and Molecular Biology Research Center; Department of Parasitology, School of Medicine, Shaheed Beheshti University of Medical Sciences, Tehran, Iran. bahram_14@yahoo.com, kazemi@sbmu.ac.ir
Bandehpour
Mojgan
M
Seyed
Negar
N
Roozbehi
Mehrnaz
M
Mosaffa
Nariman
N
D003001:Cloning, Molecular; D018076:DNA, Complementary; D004591:Electrophoresis, Polyacrylamide Gel; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D014758:Viral Core Proteins / Q000235:genetics* / Q000276:immunology
D018076:DNA, Complementary; D011994:Recombinant Proteins; D014758:Viral Core Proteins; C069130:nucleocapsid protein, Hepatitis C virus