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Reverse Genetics to Engineer Positive-Sense RNA Virus Variants

Article DOI: 10.3791/63685-v • 15:49 min • June 9th, 2022
June 9th, 2022 •
1, 1, 2, 1
1Virology Section, Department of Life Sciences, Shiv Nadar University, 2Gastroenterology, Sanjay Gandhi Postgraduate Institute of Medical Sciences

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The protocol describes a method for introducing controllable genetic diversity in the hepatitis C virus genome by combining full-length mutant RNA synthesis using error-prone PCR and reverse genetics. The method provides a model for phenotype selection and can be used for 10 kb long positive-sense RNA virus genomes.

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Reverse Genetics Positive-sense RNA Virus Variants Directed Evolution RNA Genome Error-prone PCR Viral Mutants Phenotypes Of Interest Full-length Mutant RNA Synthesis (FL-MRS) Library Genetic Diversity DNA Polymerase Fidelity Template Mutant RNA Synthesis Host Cells Non-selected Input Pool Viral-directed Evolution Hepatitis C Virus (HCV) JFH1 Isolate Adaptation Replication Viral Genes Pathogenesis Antiviral Resistance
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