Booster PCR: evaluation of an improved method for amplification of a few HIV-1 proviral DNA sequences.
A biphasic polymerase chain reaction designed as booster PCR for the detection of human immunodeficiency virus type-1 (HIV-1) was evaluated in samples containing a very low number of target DNA. We examined DNA samples obtained from chronically infected H9/HTLV-III B cells and purified plasmidic DNA containing the entire HIV-1 genome. By using booster PCR we detected HIV-1 DNA sequences up to 5 infected cells in samples containing about 2 micrograms of genomic DNA, and up to 1 copy of plasmidic DNA in samples containing about 0.5 microgram of genomic DNA. Otherwise by using standard PCR HIV-DNA up 100 infected cells and up to 20 copies from plasmidic DNA could be detected. Our experiments in amplification of HIV-1 proviral DNA have demonstrated that booster PCR enhances sensitivity of detection of standard PCR in small quantities of target sequences at least 20-fold with no loss of specificity.
Institute of Infectious Diseases, University of Pavia, IRCCS San Matteo, Italy.
G
G
Filice
M
M
Debiaggi
P M
PM
Cereda
E
E
Romero
D002460:Cell Line; D004279:DNA, Viral / Q000032:analysis*; D005069:Evaluation Studies as Topic; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D011533:Proviruses / Q000235:genetics*; D012680:Sensitivity and Specificity
D004279:DNA, Viral