Primer specific and mispair extension analysis (PSMEA) as a simple approach to fast genotyping.
A simple method, primer specific and mispair extension analysis (PSMEA) with pfu DNA polymerase was developed for genotyping. PSMEA is based on the unique properties of 3'-->5' exonuclease proofreading activity. In the presence of an incomplete set of dNTPs, pfu was found to be extremely discriminative in nucleotide incorporation and proofreading at the initiation step of DNA synthesis, completely preventing primer extension when mispair(s) are found adjacent to the 3'-end of the primer. This has allowed us to accurately detect nucleotide variations, deletions and insertions for fast genotyping.
The Canadian Red Cross Society, Ottawa, Ontario K1G 4J5, Canada, Laboratoire de Santé Publique du Québec, Ste-Anne-de-Bellevue, Quebéc H9X 3R5, Canada. yhn@redcross.ca
Y W
YW
Hu
E
E
Balaskas
G
G
Kessler
C
C
Issid
L J
LJ
Scully
D G
DG
Murphy
A
A
Rinfret
A
A
Giulivi
V
V
Scalia
P
P
Gill
D020137:Base Pair Mismatch; D001483:Base Sequence; D004247:DNA / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics; D004259:DNA-Directed DNA Polymerase; D005821:Genetic Techniques; D005838:Genotype; D016174:Hepacivirus / Q000235:genetics; D006801:Humans; D017422:Sequence Analysis, DNA; D013789:Thalassemia / Q000235:genetics
D017931:DNA Primers; D004279:DNA, Viral; D004247:DNA; C072146:Pfu DNA polymerase; D004259:DNA-Directed DNA Polymerase