title: Polymerase chain reaction detection of type D simian retrovirus proviral DNA from infected macaques.
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). Env PCR fragments were readily detected from as few as 10(4) 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.
authors.affiliation: Division of Neuroscience, Oregon Regional Primate Research Center, Beaverton 97006.
authors.forename: K Y
authors.initials: KY
authors.lastname: Pilcher
authors.forename: N
authors.initials: N
authors.lastname: Avery
authors.forename: S M
authors.initials: SM
authors.lastname: Shiigi
authors.forename: N
authors.initials: N
authors.lastname: Pangares
authors.forename: A
authors.initials: A
authors.lastname: Malley
authors.forename: M K
authors.initials: MK
authors.lastname: Axthelm
authors.forename: C A
authors.initials: CA
authors.lastname: Machida
mesh_terms: D000818:Animals; D001483:Base Sequence; D017931:DNA Primers; D004279:DNA, Viral / Q000032:analysis*; D008214:Lymphocytes / Q000821:virology*; D008251:Macaca; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D015301:Retroviruses, Simian / Q000235:genetics / Q000302:isolation & purification*; D016097:Simian Acquired Immunodeficiency Syndrome / Q000097:blood / Q000821:virology*
chemical_list: D017931:DNA Primers; D004279:DNA, Viral