title: Rapid Determination of HIV-1 Mutant Frequencies and Mutation Spectra Using an mCherry/EGFP Dual-Reporter Viral Vector.
abstract: The high mutation rate of human immunodeficiency virus type-1 (HIV-1) has been a pivotal factor in its evolutionary success as a human pathogen, driving the emergence of drug resistance, immune system escape, and invasion of distinct anatomical compartments. Extensive research has focused on understanding how various cellular and viral factors alter the rates and types of mutations produced during viral replication. Here, we describe a single-cycle dual-reporter vector assay that relies upon the detection of mutations that eliminate either expression of mCherry or enhanced green fluorescent protein (EGFP). The reporter-based method can be used to efficiently quantify changes in mutant frequencies and mutation spectra that arise due to a variety of factors, including viral mutagens, drug resistance mutations, cellular physiology, and APOBEC3 proteins.
authors.affiliation: Institute for Molecular Virology, University of Minnesota, Minneapolis, MN, 55455, USA.|Molecular, Cellular, Developmental Biology & Genetics Graduate Program, University of Minnesota, Minneapolis, MN, 55455, USA.
authors.forename: Jonathan M O
authors.initials: JM
authors.lastname: Rawson
authors.affiliation: Institute for Molecular Virology, University of Minnesota, Minneapolis, MN, 55455, USA.|Department of Diagnostic and Biological Sciences, School of Dentistry, University of Minnesota, Minneapolis, MN, 55455, USA.
authors.forename: Christine L
authors.initials: CL
authors.lastname: Clouser
authors.affiliation: Institute for Molecular Virology, University of Minnesota, Minneapolis, MN, 55455, USA. mansky@umn.edu.|Molecular, Cellular, Developmental Biology & Genetics Graduate Program, University of Minnesota, Minneapolis, MN, 55455, USA. mansky@umn.edu.|Department of Diagnostic and Biological Sciences, School of Dentistry, University of Minnesota, Minneapolis, MN, 55455, USA. mansky@umn.edu.|Department of Microbiology, University of Minnesota, Graduate Program, Mayo Mail Code 196, 1460 Mayo Building, 420 Delaware Street SE, Minneapolis, MN, 55455, USA. mansky@umn.edu.
authors.forename: Louis M
authors.initials: LM
authors.lastname: Mansky
mesh_terms: D018929:Cell Culture Techniques / Q000379:methods; D002460:Cell Line; D004279:DNA, Viral / Q000235:genetics* / Q000302:isolation & purification; D005456:Fluorescent Dyes / Q000378:metabolism; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics*; D015658:HIV Infections / Q000821:virology*; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics*; D009154:Mutation; D059645:Mutation Rate; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D017422:Sequence Analysis, DNA / Q000379:methods; D000097573:Red Fluorescent Protein
chemical_list: D004279:DNA, Viral; D005456:Fluorescent Dyes; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins; D000097573:Red Fluorescent Protein; C485184:enhanced green fluorescent protein
keywords: Diversity; Evolution; Lentivirus; Mutagenesis; Retroviral vector; Retrovirus; Reverse transcription