Thermus aquaticus DNA polymerase-catalysed chain reaction for the detection of human papillomaviruses.
To delineate the conditions for the polymerase chain reaction (PCR) using primers specific for human papillomavirus (HPV) types 6b, 16 and 18, a number of important technical features were analysed. Buffer, concentrations of magnesium, Taq polymerase, primers and DNA templates, annealing temperature, and extension time were studied by a combination of gel electrophoresis, Southern and slot-blot hybridization. Amplification of E6 gene fragments of HPV-16 and HPV-18 generated bands of 110 bp and 154 bp respectively, as predicted. However, amplification of a segment within the long control region of HPV 6b yielded an unexpected size of 340 bp. Different conditions were found for each HPV type-specific primer pair. These results, and the applications of PCR in HPV research and in an increasingly wide range of fields in medical virology are discussed.
Chow
V T
VT
Institute of Molecular and Cell Biology, National University of Singapore.
Tham
K M
KM
Bernard
H U
HU
D001483:Base Sequence; D015139:Blotting, Southern; D002460:Cell Line; D004279:DNA, Viral / Q000096:biosynthesis; D004259:DNA-Directed DNA Polymerase; D006801:Humans; D008969:Molecular Sequence Data; D021141:Nucleic Acid Amplification Techniques; D015345:Oligonucleotide Probes; D027383:Papillomaviridae / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods*; D019914:Taq Polymerase; D013696:Temperature; D013997:Time Factors; D014412:Tumor Virus Infections / Q000175:diagnosis*
D004279:DNA, Viral; D015345:Oligonucleotide Probes; D019914:Taq Polymerase; D004259:DNA-Directed DNA Polymerase