title: Modified Genomic Sequencing PCR Using the MiSeq Platform to Identify Retroviral Integration Sites.
abstract: High-throughput mapping of retroviral vector integration sites (RIS) has become an invaluable tool to evaluate novel gene therapy vectors and to track clonal contribution in preclinical and clinical studies. Beard et al. (Methods Mol Biol 2014;1185:321-344) described an improved protocol developed for efficient capture, sequencing, and analysis of RIS that preserves gene-modified clonal contribution information. Here we describe adaptations to the previously published modified genomic sequencing PCR (MGS-PCR) protocol using the Illumina MiSeq paired-end sequencing platform. Lentiviral, gammaretroviral, and foamy virus vector integrations were analyzed. MGS-PCR using the MiSeq platform allows for the use of merged paired-end reads, which allows for efficient localization of RIS to published genomes.
authors.affiliation: 1 Washington State University College of Pharmacy , Spokane, Washington.
authors.forename: Dustin T
authors.initials: DT
authors.lastname: Rae
authors.affiliation: 1 Washington State University College of Pharmacy , Spokane, Washington.
authors.forename: Casey P
authors.initials: CP
authors.lastname: Collins
authors.affiliation: 1 Washington State University College of Pharmacy , Spokane, Washington.
authors.forename: Jonah D
authors.initials: JD
authors.lastname: Hocum
authors.affiliation: 2 School of Molecular Biosciences, Washington State University , Pullman, Washington.
authors.forename: Diana L
authors.initials: DL
authors.lastname: Browning
authors.affiliation: 1 Washington State University College of Pharmacy , Spokane, Washington.|2 School of Molecular Biosciences, Washington State University , Pullman, Washington.
authors.forename: Grant D
authors.initials: GD
authors.lastname: Trobridge
mesh_terms: D000818:Animals; D056905:Chromosome Breakpoints; D019295:Computational Biology / Q000379:methods; D005822:Genetic Vectors / Q000235:genetics; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D016133:Polymerase Chain Reaction; D012190:Retroviridae / Q000145:classification / Q000502:physiology*; D017422:Sequence Analysis, DNA / Q000379:methods; D016662:Virus Integration