Ідентифікація алельних варіантів мікросателітної ДНК веслоноса (Polyodon Spathula)

Анотація

IDENTIFICATION OF ALLELIC VARIANTS OF PADDLEFISH (POLYODON SPATHULA) MICROSATELLITE DNA

Kh. Kurta, O. Malysheva, A. Getya, B. Grishyn, L. Shynkarenko, 
V. Spyrydonov

In modern conditions of the development of domestic aquaculture, more and more fish farms in Ukraine are aimed at increasing the production of valuable fish products both for commercial realization in the domestic market and for export. Nowadays, such valuable fish products are obtained from representatives of Acipenseriformes species, including paddlefish (Polyodon spathula). The intensive economic use of this object is due to its biological characteristics and valuable economic properties: the ability to live in fresh water, the way of feeding on plankton, rapid growth rates, high ecological plasticity to different growing conditions, meat quality and delicious caviar.

Currently, an increase in the production of commercial products of the paddlefish requires a substantial expansion of its broodstock within the farms. Therefore, the question of controlling and maintaining the genetic diversity of broodstock of this species of fish is a priority for fish farms, which are aimed at increasing of artificial reproduction and the productivity of aquaculture.

The ukrainian populations of paddlefish were formed in the 1960's by introducing a limited number of ichthyological material from the United States and Russia. Rearing and breeding of the limited quantities of ichthyologic material can lead to low genetic diversity and high probability of inbreeding depression among offspring. As a result, it can lead to a gradual decline in productive qualities, decrease in fish resistance to diseases and resistance to the effects of adverse external environmental factors.

One of the important parts of effective breeding in the formation of broodstock is the complex assessment of the genetic diversity of the breeding groups of fish. Under such conditions, there is a need for the use of modern methods of monitoring genetic processes occurring in artificial populations of paddlefish.

In recent years, population-genetic studies have become widely used in molecular genetic methods that allow to analyze microsatellite DNA of living organisms. Microsatellites are highly polymorphic and effective DNA markers that are used to evaluate the genetic structure of populations and allow to investigate paternal relationships between individuals. In addition, microsatellite DNA markers can be used for individual identification, which allows to see changes in the genotype of each individual and compare them among themselves. Despite the high environmental adaptability, the molecular evolution velocity of nuclear DNA of Acipenseriformes species is rather low, which allows the use of microsatellite DNA markers to study the genetic features of all representatives of this order.

Thus, the use of microsatellite DNA analysis will provide effective genetic monitoring of broodstock and their rational use for improving the reproduction and rearing of aquaculture objects.

Therefore, the purpose of this study was to investigate the genetic structure and identify allelic variants of microsatellite DNA in the artificial population of paddlefish (Polyodon spathula).

According to results of microsatellite DNA analysis of the genetic structure of the studied group of paddlefish, 24 allelic variants were identified for each locus. Psp26 and Psp28 loci were the most polymorphic (7 allelic variants), and the Psp21 locus was the least polymorphic (4 allelic variants). For the Psp12 locus, two new allele variants (214 and 216 bp) were identified, and for the Psp26 loci, one new allele variant (164 bp) was identified. The mean number of alleles per locus (Na) was 6,0 and ranged from 4 (Psp21) to 7 (Psp26, Psp28).

The mean values of the observed (Ho) and expected (He) heterozygosity were at 0,671 and 0,599, respectively. The maximum value of Ho was for the Psp28 locus - 1,000, the minimum (0,395) for the Psp21 locus. The expected heterozygosity (He) ranged from 0,411 to 0,685 for Psp21 and Psp26 loci, respectively.

In addition, we established a reliable (p <0.05) shortage of heterozygous genotypes at the Psp26 locus, while, on the contrary, a significant surplus of heterozygous genotypes (p <0.01) was  revealed at the Psp28 locus. This means that the studied fish group is currently genetically balanced.

The mean value of index of polymorphism for the studied DNA loci was at the level of 0,548, indicating a sufficient level of polymorphism for the selected DNA-markers for this species of fish (PIC> 0,500).

The probability of excluding (PE) varied from 0,111 to 1,000 for Psp21 and Psp28 loci respectively, and means value was at 0,452. The value of the combined probability of excluding (CPE) was at 1,000, indicating a high level of informativeness of the selected panel of microsatellite DNA markers.

Index of polymorphism (PIC) for the studied loci for paddlefish ranged from 0,361 for Psp21 locus to 0,664 for the Psp26 locus, indicating that the Psp21 locus was least polymorphic, and the Psp26 locus was the most polymorphic. The mean value of the index of polymorphism for the studied DNA loci was 0,548, indicating a sufficient level of polymorphism for the selected markers for this species (PIC> 0,500).

The probability of excluding (PE) averaged 0,452 and ranged from 0,111 (Psp21) to 1,000 (Psp28). The high value of PE for the Psp28 (1,000) locus was due to the presence of only heterozygous genotypes in the 100% studied individuals of paddlefish for this DNA marker.

According to population-genetic calculations it was established that the use of the selected panel of microsatellite DNA markers allows to genotype the paddlefish with a sufficiently high level of informativity.

Thus, the results of microsatellite DNA studies allowed detecting and evaluating the changes that occurred in the genetic structure of paddlefish. The identified specific allelic variants will allow identification of existing local groups of this species, which can be used to monitor the genetic status and effectiveness of biodiversity conservation of artificial populations in modern fish farms of Ukraine.