title: GeXPS multiplex PCR analysis of the simian varicella virus transcriptome in productively infected cells in culture and acutely infected ganglia.
abstract: Simian varicella zoster virus (SVV) infection of non-human primates serves as a model to study varicella zoster virus (VZV) infection and pathogenesis in humans. While macroarray analysis detected all 69 predicted unique open reading frames (ORFs) in SVV-infected cells in culture, it lacked the sensitivity to detect the low-abundance transcripts expressed in latently infected monkey ganglia. Recently, a multiplex RT-PCR assay using the GenomeLab Genetic Analysis System (GeXPS) identified 10 VZV transcripts in latently-infected human ganglia. GeXPS was used to analyze the SVV transcriptome in SVV-infected monkey cells in culture as well as in acutely infected ganglia from African green monkeys. Oligonucleotide primers containing both SVV ORF- and cell-specific sequences linked to universal DNA sequences were used in RT-PCR to produce products of predetermined sizes. Amplification products were resolved by capillary gel electrophoresis and detected by fluorescence spectrophotometry. Transcripts corresponding to all 69 predicted SVV ORFs, in addition to transcripts within the leftward end region and ORF 61 antisense transcripts were detected in virus-infected cells in culture. Except for two transcripts (ORFs 14 and 35), all transcripts found in infected tissue culture cells were also found in acutely infected monkey ganglia.
authors.affiliation: Department of Neurology, University of Colorado Denver School of Medicine, 12700 East 19th Avenue, Mail Stop B182, Aurora, CO 80045, USA. Maria.Nagel@ucdenver.edu
authors.forename: Maria A
authors.initials: MA
authors.lastname: Nagel
authors.forename: Alexander
authors.initials: A
authors.lastname: Choe
authors.forename: Don
authors.initials: D
authors.lastname: Gilden
authors.forename: Vicki
authors.initials: V
authors.lastname: Traina-Dorge
authors.forename: Randall J
authors.initials: RJ
authors.lastname: Cohrs
authors.forename: Ravi
authors.initials: R
authors.lastname: Mahalingam
mesh_terms: D000818:Animals; D002460:Cell Line; D002522:Chlorocebus aethiops; D019075:Electrophoresis, Capillary; D005470:Fluorometry; D005724:Ganglia / Q000821:virology*; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D016366:Open Reading Frames; D059467:Transcriptome; D018140:Varicellovirus / Q000235:genetics* / Q000254:growth & development; D014773:Virology / Q000379:methods
keywords: GeXPS; PCR; Simian varicella virus transcriptome