Development of a set of oligonucleotides for the identification of the Sakhalin sturgeon Acipenser mikadoi Hilgendorf, 1892 by PCR

Sturgeons (family Acipenseridae) are valuable commercial fish and aquaculture resources. The Acipenser genus includes about 20 species, 12 of which are recorded for the Russian territory. One of the rarest is the Sakhalin sturgeon A. mikadoi Hilgendorf, 1892. At present, its population size is significantly decreased and the species is close to extinction. Natural populations of the species have survived in the Tumnin River in Khabarovsk Region and the Viakhtu River in Sakhalin Region. Vihtu River in Sakhalin Region. Due to the small numbers of rare and endangered fish species, noninvasive approaches, including the use of DNA from the environment, appear to be the most suitable for monitoring their species and genetic diversity. The method is well established in this area and has been successfully tested for the monitoring of several species of rare and endangered sturgeons. This paper presents the results of the development of primers specific for the mitochondrial DNA of the species A. mikadoi.

A matrix of 15 sequences of complete mitochondrial genomes from 3 sturgeon species whose ranges may overlap - A. mikadoi (KX276658), A. dauricus (KJ402277) and A. schrenckii (MH973728- MH973734, KX276660, KX276659, KF150287, KC905169, KC820796) - was generated. A. medirostris (NC_028405), which is genetically closest to the Sakhalin sturgeon, was also included in the analysis. A sliding window algorithm was applied to the sequence matrix. A graph showing the distribution of divergence values along the mitochondrial genome of sturgeon was plotted based on the results of the analysis Fig. 1.

Based on matrices of individual genome fragments (16S rRNA, COI, ND1, and D-loop), a search for species-specific primers was performed in the DECIPHER program. A Primer-BLAST algorithm was run based on each pair of primers. Taxa Actinopterygii and Acipenseridae were used as references separately in different runs. In addition, primers were checked using local reference sequences of the complete mitochondrial genome of representatives of the genus Acipenser. The results of primer evaluation are summarized in the table Table 1. As a result, a combination of primer pairs developed on the basis of D-loop and COI fragments seems to be the most successful for error-free identification of Sakhalin sturgeon.