A recombinant replication-competent hepatitis C virus expressing Azami-Green, a bright green-emitting fluorescent protein, suitable for visualization of infected cells.
نویسندگان
چکیده
The hepatitis C virus (HCV) production system consists of transfecting the human hepatoma cell line Huh7 with genomic HCV RNA (JFH1). To monitor HCV replication by fluorescence microscopy, we constructed a recombinant HCV clone expressing Azami-Green (mAG), a bright green fluorescent protein, by inserting the mAG gene into the nonstructural protein 5A (NS5A) gene; the resultant clone was designated JFH1-hmAG. The Huh-7.5.1 (a subclone of Huh7) cells transfected with JFH1-hmAG RNA were found to produce cytoplasmic NS5A-mAG, as readily visualized by fluorescence microscopy, and infectious virus, as assayed with the culture supernatant, indicating that JFH1-hmAG is infectious and replication-competent. Furthermore, the replication of this virus was inhibited by interferon alpha in a dose-dependent manner. These results suggest that JFH1-hmAG is useful for studying HCV life cycle and the mechanism of interferon's anti-HCV action and for screening and testing new anti-HCV drugs.
منابع مشابه
Transient expression of green fluorescent protein in radish (Raphanus sativus) using a turnip mosaic virus based vector
It is possible to use transgenic plants, as bioreactors, for the production of recombinant inexpensive chemicals and drug components. Transient gene expression is an appropriate alternative to stable transformation because it allows an inexpensive and rapid method for expression of recombinant proteins in plant tissues. In recent years, plant viral vectors have been increasingly developed as su...
متن کاملHepatitis C virus p7 protein is localized in the endoplasmic reticulum when it is encoded by a replication-competent genome.
p7 protein is a small protein encoded by Hepatitis C virus (HCV) that functions as an ion channel in planar lipid bilayers. The function of p7 is vital for the virus life cycle. In this study, the p7 protein of genotype 2a (strain JFH1; the only strain that replicates and produces virus progeny in vitro) was tagged with either an enhanced green fluorescent protein (eGFP) or a haemagglutinin (HA...
متن کاملEstablishment of NS3 tumor cell line expressing Hepatitis C virus Non-structural Protein 3 as valuable tool for HCV challenging in mice
Introduction: Hepatitis C virus (HCV) is one of the major medical problems. Human and chimpanzees are the only specific hosts which are naturally susceptible to HCV infection. Mice and other common laboratory animals are resistant to the virus, hence HCV prophylactic and therapeutic researches are very difficult and challenging. HCV non-structural protein 3 (NS3) is one of the most attractive t...
متن کاملA Gaussia Luciferase Cell-Based System to Assess the Infection of Cell Culture- and Serum-Derived Hepatitis C Virus
Robust replication of hepatitis C virus (HCV) in cell culture occurs only with the JFH-1 (genotype 2a) recombinant genome. The aim of this study was to develop a system for HCV infection quantification analysis and apply it for the selection of patient sera that may contain cell culture infectious viruses, particularly of the most clinically important genotype 1. Initially, a hepatoma cell line...
متن کاملDevelopment of an infectious surrogate hepatitis C virus based on a recombinant vesicular stomatitis virus expressing hepatitis C virus envelope glycoproteins and green fluorescent protein.
To develop surrogate viruses for hepatitis C virus (HCV), we previously produced recombinant vesicular stomatitis viruses (rVSVs) lacking glycoprotein G but instead expressing chimeric HCV E1/E2 fused to G. These rVSVs were not infectious in HCV-susceptible hepatoma cells. In this study, to develop an infectious surrogate HCV based on an rVSV (vesicular stomatitis virus [VSV]/HCV), we generated...
متن کاملذخیره در منابع من
با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید
برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید
ثبت ناماگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید
ورودعنوان ژورنال:
- Biochemical and biophysical research communications
دوره 377 1 شماره
صفحات -
تاریخ انتشار 2008