Moloney murine leukemia reverse transcriptase suspect in the production of multiple misincorporations during hprt cDNA synthesis.
Our laboratory has characterized several hundred mutant hprt cDNAs produced using Moloney murine leukemia reverse transcriptase to convert mRNA to cDNA. During the characterization of these mutants we have detected six T-lymphocyte mutants that demonstrate multiple G:C --> A:T transitions along the hprt cDNA coding sequence. Attempts to repeat the mRNA to cDNA conversion and subsequent characterization have demonstrated that the multiple transitions are likely artifacts. We suggest that reverse transcriptase is directly responsible for these multiple base substitutions and as such, that multiple mutations be viewed as suspect requiring confirmation at the genomic level.
Curry
J
J
Department of Biology, University of Victoria, British Columbia, Canada. jcurry@uvic.ca
Glickman
B W
BW
D000818:Animals; D004261:DNA Replication; D018076:DNA, Complementary / Q000096:biosynthesis; D007041:Hypoxanthine Phosphoribosyltransferase / Q000235:genetics*; D066298:In Vitro Techniques; D051379:Mice; D008979:Moloney murine leukemia virus / Q000201:enzymology*; D016296:Mutagenesis; D009692:Nucleic Acid Heteroduplexes; D016133:Polymerase Chain Reaction; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism*
D018076:DNA, Complementary; D009692:Nucleic Acid Heteroduplexes; D007041:Hypoxanthine Phosphoribosyltransferase; D012194:RNA-Directed DNA Polymerase