ISSN 1022-7954, Russian Journal of Genetics, 2006, Vol. 42, No. 1, pp. 16–26. © Pleiades Publishing, Inc., 2006.
Original Russian Text © I.A. Koﬁadi, D.V. Rebrikov, 2006, published in Genetika, 2006, Vol. 42, No. 1, pp. 22–32.
Single nucleotide polymorphisms (SNPs) are the
variable positions in DNA sequence. Scientists agreed
to consider as polymorphisms only those DNA sites for
which the frequency of the least prevalent variant
amounts to less than 1% [1–4]. SNP is the most fre-
quent reason underlying existence of several variants of
the same gene (alleles); the vast majority of variants in
the human genome result from SNPs [5, 6]. SNPs are
unevenly distributed in the genome; however, 1000
nucleotides on average carry one variable nucleotide
[7–10]. Thus, two unrelated human genomes differ in
approximately three million SNPs.
Each individual polymorphism in the population
may be represented by two, three, or four variants
(according to the number of possible nucleotides in a
DNA position); however, the majority of SNPs exist in
two variants (are biallelic) [1, 11, 12]. Presumably, the
majority of SNPs have originated due to mutations
ﬁxed in the population . The most frequent polymor-
phism is variation of bases of the same type (purines or
pyrimidines). The polymorphisms with purine/pyrimi-
dine alleles are met essentially rarer. The frequencies of
SNPs of various types in the human genome amount to
63% for A/G, 17% for A/C, 8% for G/C, and 4% for A/T.
Insertions and deletions form the rest 8% .
The polymorphisms localized to the genome coding
regions are of most practical interest. The public data-
bases now contain information about two million
SNPs; however, only a small part of them (approxi-
mately 1%) are located in transcribed regions and may
inﬂuence the structure and function of the product
encoded by the corresponding gene .
The SNPs located in coding regions may be associ-
ated with certain phenotypes. Recently, research into
detection of polymorphisms connected with individual
propensity for diseases determined by biochemical pro-
ﬁle, tolerance to environmental effects (in particular,
various medical drugs), etc. [14–19] became wide-
spread. The databases, such as dbSNP of the National
Center for Biomedical Information, United States
(http://www.ncbi.nlm.nih.gov/SNP/) and ENSEMBL
of the European Molecular Biology Laboratory, Ger-
many (http://www.ensembl.org), contain public data
about oligonucleotide polymorphisms in the human
genome. The main sources of data on new SNPs for the
public databases to date are the SNP Consortium
(TSC), the Sanger Center (UK), and Washington Uni-
versity (United States) .
A high density and evolutionary stability of SNPs
make them one of the most attractive genetic markers
[11, 21–23]. SNP analysis is widely used in population
studies and genetic mapping [3, 12].
All these facts explain a great diversity of methods
for SNP detection that appeared recently, and it is fre-
quently a complicated issue for a researcher to decide
on a particular approach. When developing various
methods, the authors use various basic principles, such
as allele-speciﬁc PCR, hybridization with an oligonu-
cleotide probe, various variants of DNA sequencing,
enzymatic cleavage, etc. [24–32].
In this work, we attempted to systematize and
brieﬂy describe the state-of-the-art methods for SNP
detection that are based on allele-speciﬁc PCR and
hybridization with oligonucleotide probe without using
additional enzymes, such as ligases, restriction endonu-
cleases, exonucleases, etc. (except for 5'-exonuclease
activity of nonrecombinant
polymerase). In the
ﬁnal part of the paper, we characterized the considered
methods according to three criteria: their accuracy,
cost, and simplicity.
Methods for Detecting Single Nucleotide Polymorphisms:
Allele-Specific PCR and Hybridization
with Oligonucleotide Probe
I. A. Kofiadi and D. V. Rebrikov
DNA-Technology, Moscow, 115478 Russia; fax (095) 116-49-02; e-mail: email@example.com
Received July 19, 2005
—The existing diversity of the methods for detecting single nucleotide polymorphisms is so great that
may perplex an unsophisticated researcher who chooses the appropriate molecular genetic toolkit. In this work,
we tried to systematize and brieﬂy describe the state-of-the-art methods for detecting oligonucleotide polymor-
phisms that are based on allele-speciﬁc PCR and hybridization with oligonucleotide probe as well as to char-
acterize the methods considered with respect to their accuracy, cost, and simplicity.