Document 0166
 DOCN  M95B0166
 TI    Increased reliability of selective PCR by using additionally mutated
       primers and a commercial Taq DNA polymerase enhancer.
 DT    9511
 AU    De Milito A; Catucci M; Iannelli F; Romano L; Zazzi M; Valensin PE;
       Dipartimento di Biologia Molecolare, Universita di Siena, Italy.
 SO    Mol Biotechnol. 1995 Apr;3(2):166-9. Unique Identifier : AIDSLINE
       MED/95346400
 AB    A reliable selective PCR procedure that combines the use of additionally
       mutated primers with the specificity-enhancing properties of a
       commercial preparation (Perfect Match, Stratagene) is described. The
       human immunodeficiency virus type 1 pol gene point mutations known to
       confer in vitro resistance to azidothymidine were examined as a model
       for optimization of the assay. The usual strategy of deliberately
       introducing an additional mismatch 1 residue from the 3' end in the
       wild-type and mutant primers did not allow reproducible discrimination
       between wild-type and mutant target sequences. Addition of minimal
       amounts of Perfect Match to the same PCR mixtures resulted in a
       significantly enlarged range of selective annealing temperatures,
       providing a valuable and cost-effective means for reliable detection of
       known mutations by selective PCR.
 DE    Base Sequence  Drug Resistance, Microbial  DNA/*ISOLATION & PURIF  DNA
       Polymerases/CHEMISTRY  DNA Primers/GENETICS  Genes, pol  Human
       HIV-1/DRUG EFFECTS/GENETICS  Molecular Sequence Data  *Point Mutation
       Polymerase Chain Reaction/*METHODS  Reproducibility of Results  Support,
       Non-U.S. Gov't  Temperature  Zidovudine/PHARMACOLOGY  JOURNAL ARTICLE

       SOURCE: National Library of Medicine.  NOTICE: This material may be
       protected by Copyright Law (Title 17, U.S.Code).