Polymerase chain reaction detection of type D simian retrovirus proviral DNA from infected macaques

Kirsten Y. Pilcher, Nancy Avery, Stanley M. Shiigi, Nancee Pangares, Arthur Malley, Michael K. Axthelm, Curtis A. Machida

Research output: Contribution to journalArticle

6 Scopus citations

Abstract

A simple polymerase chain reaction (PCR) approach was developed for detection of Type D simian retrovirus (SRV) serogroup 2 proviral DNA using peripheral blood lymphocytes (PBLs) obtained from infected macaques. PCR primer pairs were developed against serogroup 2 envelope (env) gene sequence, and fidelity of PCR fragment amplification was determined using molecularly cloned SRV serogroup 2 (D2/RHE/OR) DNA, and genomic DNA from Raji cells independently infected with different SRV serogroups. One primer pair exhibiting high fidelity was then utilized for PCR detection of serogroup 2 proviral DNA from PBLs, and from cells sorted into immune cell subpopulations by fluorescent-activated cell sorting (FACS). Enυ PCR fragments were readily detected from as few as 104 PBLs or immune cell subpopulations. In addition, highly specific PCR primers against serogroups 1 and 3 were utilized to detect proviral DNA from Raji cells infected with SRV serogroups. In all cases, primers designed to amplify serogroups 1,2, and 3 proviral DNA were specific for their intended serogroup. This primer information and development of a PCR approach for detection of specific SRV proviral DNA will be of potential utility as a rapid surveillance tool in monitoring type D simian retrovirus infection within Asian macaque colonies.

Original languageEnglish (US)
Pages (from-to)75-86
Number of pages12
JournalJournal of Virological Methods
Volume50
Issue number1-3
DOIs
StatePublished - Dec 1994

Keywords

  • Immune cell subpopulation
  • Peripheral blood lymphocyte
  • Polymerase chain reaction
  • Type D simian retrovirus

ASJC Scopus subject areas

  • Virology

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