New Multiplex PCR Method for Identification of East European Green Frog Species and Their Hybrids

O. Ermakov, A. Ivanov, S. Titov, A. Svinin, S. N. Litvinchuk


A molecular multiplex PCR method for identification of East European green frog species (Pelophylax ridibundus, P. cf. bedriagae and P. lessonae) and their hybrids was developed. This simple and rapid method can be used for identification of species-specific mitochondrial and nuclear DNA. The method is based on species-specific differences in primary structure of the subunit 1 of the mitochondrial cytochrome C oxidase gene (COI) and the intron-1 of the nuclear serum albumin gene (SAI-1). Based on the method, we analyzed numerous individuals of these species and their hybrids from East European Plain, the Crimea, the Caucasus, the Ural, as well as introduced populations from Western Siberia and the Kamchatka. In all cases, identification of species performed by use of the multiplex PCR method coincided with results of study of primary nucleotide sequences.


Pelophylax ridibundus; Pelophylax cf. bedriagae; Pelophylax lessonae; Pelophylax esculentus; rapid species identification; water frogs; cytochrome C oxidase; serum albumin

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