ISSN 2415-8860 (Online), ISSN 0372-4123 (Print)
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Ukr. Bot. J. 2018, 75(6): 576–584
Plant Physiology, Biochemistry, Cell and Molecular Biology

Study of intron length polymorphism of the α-tubulin genes as a method of analysis of the genetic differentiation in plants

Pirko Ya.V., Postovoitova A.S., Rabokon A.M., Kalafat L.O., Privalikhin S.M., Bilonozhko Yu.O., Pirko N.M., Blume Ya.B.

A new type of DNA markers based on the analysis of the α-tubulin 1st intron polymorphism has been developed and implemented. The bioinformatics search for α-tubulin genes of Arabidopsis thaliana, Linum usitatissimum, Oryza sativa, Solanum tuberosum, and S. lycopersicum has been carried out. It has been shown that most genes of α-tubulin contained 4-5 exons and 3 to 4 introns. Several exceptions have been identified, including the A. thaliana α-tubulin gene TUBA6 that contained only 2 exons and 1 intron, and the TUBA4 gene that consisted of 3 exons and 2 introns. It have been established that the lengths of the introns varies considerably, even within the same species. A certain systemicity was found in the number of nucleotide pairs of exons. Based on data on the analysis of the α-tubulin gene exon-intron structure, the pair of universal degenerate primers have been created and the evaluation of the 1st intron length polymorphism of α-tubulin genes in Arabidopsis thaliana and various varieties of L. usitatissimum, O. sativa, S. tuberosum, S. lycopersicum have been studied. It was shown the formation of species-specific DNA profiles that contained a different number of first intron amplicons of the α-tubulin genes. The range of the size variation of the amplicons of intron fragments, for example in S. tuberosum, was within 150 bp–2000 bp. The nature of the appearance of large DNA fragments (more than 1500 bp) in the electrophoretic spectra of the analyzed species requires additional research, since such fragments are not generally envisaged by the results of the bioinformatics analysis. The polymorphism of the length of individual fragments of α-tubulin introns among L. usitatissimum, O. sativa, S. lycopersicum, S. tuberosum varieties has been determined, which allowed to differentiate them among themselves. In general, data was obtained confirming the feasibility of further using the polymorphism of the lengths of the 1st intron of α-tubulin genes to genotyping and assessing the genetic diversity of different species (varieties) of higher plants. The developed DNA marker system is versatile and combines the reliability, speed of obtaining raw data and the simplicity of their analysis.

Keywords: DNA-marker, genotyping, polymerase chain reaction, Arabidopsis thaliana, Linum usitatissimum, Oryza sativa, Solanum tuberosum, Solanum lycopersicum

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