Viral Load Analysis of Hepatitis C Virus in Huh7.5 Cell Culture System.
BACKGROUND
Previous studies using cell culture systems for the replication of hepatitis C virus have opened new research dimensions, and paved the ways for further and detailed studies of the virus in vitro.


OBJECTIVE
The purpose of the present study was to cultivate hepatitis C virus in a cell culture system and evaluate viral amplification.


METHODS
In order to propagate hepatitis C virus, cloned whole genome of virus, JFH-1, was used. JFH-1 cDNA was introduced into strain JM109 of Escherichia coli and plasmid, containing the viral genome was purified from transformed bacteria. After XbaI digestion, RNA synthesis was induced using T7 RNA polymerase enzyme. Next, eukaryotic cell line Huh 7.5 was transfected by the purified RNA. Finally, Huh-7.5 cell line was infected with replicated virus and viral load was determined using real-time PCR (Polymerase Chain Reaction).


RESULTS
The amount of viral load, which was measured using real-time PCR was 17592 IU/mL.


CONCLUSIONS
In the present study, using cell culture, a high titer (in acceptable range) of infectious hepatitis C virus was produced. This method could be used in future studies.
Antimicrobial Resistance Research Center, Mashhad University of Medical Sciences, Mashhad, IR Iran.
Roghayeh
R
Teimourpour
Antimicrobial Resistance Research Center, Mashhad University of Medical Sciences, Mashhad, IR Iran.
Zahra
Z
Meshkat
Department of Modern Sciences and Technologies, School of Medicine, Mashhad University of Medical Sciences, Mashhad, IR Iran.
Aida
A
Gholoubi
Antimicrobial Resistance Research Center, Mashhad University of Medical Sciences, Mashhad, IR Iran.
Hosein
H
Nomani
Department of Biology, Faculty of Sciences, Ferdowsi University of Mashhad, Mashhad, IR Iran.
Sina
S
Rostami
DNA, Complementary; Hepatitis C Virus; Huh 7.5; JFH-1; Real-Time Polymerase Chain Reaction