1022-7954/01/3710- $25.00 © 2001
Russian Journal of Genetics, Vol. 37, No. 10, 2001, pp. 1130–1133. Translated from Genetika, Vol. 37, No. 10, 2001, pp. 1348–1352.
Original Russian Text Copyright © 2001 by Sidorenko, Berezovskaya.
Analysis and quantitative evaluation of genetic
diversity is one of the key problem of evolutionary biol-
ogy. Methods based on molecular DNA markers are the
powerful tool for solving this problem. The earliest and
the most widely used of these methods is analysis of
restriction fragment length polymorphism (RFLP) [1–4].
This approach consists in cleavage of the full-size
genomic DNA with restriction endonucleases followed
by separation of the obtained fragments by length and
detection of a deﬁnite sequence in these fragments with
a radioactively labeled DNA probe. As a rule, localiza-
tion and function of the detected sequence are known.
Due to variation in restriction sites in homologous
sequences, genetic distances between species and their
phylogenetic relationships can be determined. This
method can be used for assessing genetic diversity, but
its possibilities in this respect are limited because spe-
ciﬁc probes, i.e., identiﬁed and cloned DNA sequences,
are available for only a restricted number of loci, poly-
morphism of which can be determined.
Using multilocus probes, i.e., minisatellite  and
microsatellite sequences  extended the capabilities
of RFLP analysis in evaluation of genetic polymor-
phism at the DNA level. These repetitive sequences are
dispersed throughout the genome, and numerous vari-
able markers can be revealed with the corresponding
probes. Although these markers are not characterized in
detail, they are successfully used in population genetics
to identify genotypes .
In recent years, techniques based on the polymerase
chain reaction became of general use. Among these, the
RAPD analysis is the most rapid and simple . This
method involves ampliﬁcation of genomic DNA frag-
ments using a short oligonucleotide primer having an
arbitrary sequence. Numerous ampliﬁed DNA frag-
ments are then separated by electrophoresis to be tested
for polymorphism. PCR-based methods have several
advantages over RFLP, including genomic ﬁngerprint-
ing: they are rapid, require far less material, and can be
applied to collection samples. Although the structure of
RAPD markers is unknown which is the most serious
drawback of this method, this circumstance does not
interfere with a complex characteristic of genetic pro-
cesses in the total population, and, if required, can be
overcome by sequencing of the markers of interest.
RAPD markers were successfully used in popula-
tion genetics for detecting variation and differentiation
in natural populations and for determining the popula-
tion genetic structure [9–11). We have previously used
RAPD–PCR to estimate genetic polymorphism in pop-
ulations of Colorado potato beetle
In this study, individual variation of RAPD charac-
teristics was determined in the early spring generation
of a natural population of Colorado potato beetle.
MATERIALS AND METHODS
Beetles were collected in May 1999 on a locality
from Fastovskii rayon, Kievskaya oblast'. Only beetles
whose sex was unambiguously determined from the
morphological criteria described in  were used in
DNA was isolated from sterilized beetle legs
homogenized in a lysing buffer. After phenol–chloro-
form extraction DNA was precipitated by ethanol
according to a standart procedure .
The following primers were used in RAPD–PCR
analysis: P1 (GTTGCGATCC), P2 (GTTTCGCTCC),
and P4 (GTCCACACGG). The polymerase chain reac-
tion was conducted in 25
l of a buffer containing
Individual Polymorphism for RAPD Markers
in Spring Generation of Colorado Potato Beetle
A. P. Sidorenko
and O. P. Berezovskaya
Institute of Agroecology and Biotechnology, Ukrainian Agricultural Academy, Kiev, 03143 Ukraine
Schmalhausen Institute of Zoology, National Academy of Sciences of Ukraine, Kiev, 01601 Ukraine;
Received February 23, 2001
—Genetic diversity of Colorado potato beetle in a natural population from Fastovskii rayon, Kiev
oblast was estimated using RAPD. The level of polymorphism for RAPD markers was far higher in the spring
generation than in the sample obtained from the same locality in summer of the preceding year. The frequency
of individual RAPD markers was found to differ in males and females of potato beetle.