SNP genotyping by allele-specific PCR using ENA primers.
When we placed a 2'-O,4'-C-ethylene nucleic acid (ENA) residue into primers at the 3' end, or the n-1, n-2 or n-3 position, which included a single nucleotide polymorphism (SNP) site at the 3' end, only primers containing the ENA residue at the n-2 position were read by Taq DNA polymerase for amplification. When we compared n-2-position-modified ENA with the corresponding unmodified DNA primers that are often used for allele-specific PCR (AS-PCR), a greater discrimination of the SNP site by ENA primers was observed. This improvement is probably due to the difficulty of incorporating a nucleotide into a mismatched ENA primer by Taq DNA polymerase in the primer-template duplex. These results demonstrate that ENA primer-based AS-PCR would enable a rapid and reliable technique for SNP genotyping.
Koizumi
Makoto
M
Lead Discovery Research Laboratories, Sankyo Co., Ltd., 1-2-58, Hiromachi, Shinagawa-ku, Tokyo 140-8710, Japan.
Morita
Koji
K
Takagi
Miho
M
Yasumo
Hiroaki
H
Kasuya
Atsushi
A
D000483:Alleles; D017931:DNA Primers / Q000737:chemistry*; D005030:Ethylenes / Q000737:chemistry*; D005838:Genotype; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D020641:Polymorphism, Single Nucleotide; D011516:Prothrombin / Q000235:genetics
D017931:DNA Primers; D005030:Ethylenes; D011516:Prothrombin