UDC 636.1/.2:599.723: 577.2::575.174.015.3
doi: 10.15389/agrobiology.2014.4.42eng
ISSR-PCR MARKERS AND MOBILE GENETIC ELEMENTS IN HORSE (Equus caballus) GENOME
N.V. Bardukov1, A.V. Feofilov1, T.T. Glazko1, 2, V.I. Glazko1, 2
1K.A. Timiryazev Russian State Agrarian University,
49, ul. Timiryazeva, Moscow, 127550 Russia,
e-mail vglazko@yahoo.com;
2Center for Experimental Embryology and Reproductive Biotechnology,
str. 4, 12, ul. Kostyakova, Moscow, 127422 Russia,
e-mail vglazko@yahoo.com
Received March 31, 2014
The patterns and causes of the differences in the polymorphism of individual microsatellites, and species specificity of this variability are still not enough studied. Test systems developed by the International Association of Animal Genetics (ISAG) for genetic certification of agricultural animals, assessment of interbreed relations, the level of animal group consolidation, include a small number of genetic markers. The use of microsatellite loci fragments as PCR primers for polylocus genotyping and genome scan of agricultural animal species is more simple and effective methodical approach. We proposed a method for identifying positioning of microsatellite inverted repeat areas in genome sequences and for comparisons of this calculated areas to experimental DNA spectra amplified in PCR with the use of microsatellite loci as primers (ISSR-PCR markers). Here we report the results of the comparisons carried out for domestic horses (Equus caballus) of Altai breed (n = 96), Trotter breed (n = 48), Karachai breed (n = 34). The DNA fragments (99 sites) in the sequenced genome of the domestic horse, flanked by inverted repeat (GA)9C, were revealed and the presence of homologous sequences to mobile genetic elements was analyzed. The nucleotide sequences were examined in the DNA fragments (expected size/obtained size) of different location: 1250-1350/1326 bp (chromosome 17), 1250-1350/1302 bp (chromosome 2), 1250-350/1296 bp (Х-chromosome), 1250-1350/1287 bp (chromosome 20), 980/986 bp (chromosome 23), 800-900/900 bp (chromosome 24), 800-900/876 bp (Х-chromosome), 800-900/871 bp (chromosome 18), 800-900/857 bp (chromosome 3), 800-900/832 bp (chromosome 6), 800-900/823 bp (chromosome 19), 740/739 bp (chromosome 16), 580/585 bp (chromosome 1), 550/637 bp (Х-chromosome), 550/572 bp (chromosome 10), 500/494 bp (chromosome 1), 490/489 bp (chromosome 28), 380/378 bp (chromosome 11), 380/377 bp (chromosome 21), 360 (370)/364 bp (chromosome 5), 310/309 bp (chromosome 5). The PIC (polymorphic information content) values for loci were calculated. In the representatives of several breeds, the conservative and polymorphic DNA fragments were observed in the amplification product spectra obtained in the polymerase chain reaction with a primer sequence (GA)9C. It was turned out that the polymorphous DNA fragments in different animals, in most cases, contained the homologous sequences to fragments of retrotransposons (to ERV III, particularly), while the conservative ones contained homologous sequences to DNA transposons. An increased density of nucleotide sequences, potentially prone to the G4 quadruplex formation, was observed in the studied horse genomic DNA fragments, flanked by the inverted repeat (GA)9C, that could be associated with an increased recombination in these areas. A secondary structure in the DNA regions flanked by a microsatellite repeat can influence the results of genotyping in case the ISSR-PCR markers are used. The data obtained suggest the relationship between the polymorphism of DNA fragments, flanked by inverted repeats of microsatellites, and a mobility of retrotransposons.
Keywords: ISSR-PCR, anonymous DNA markers, Equus caballus, DNA transposons, retrotransposons, noncanonical DNA structures.
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