Sel’skokhozyaistvennaya Biologiya [Agricultural Biology], 2015, V. 50, № 6, pp. 756-765.

UDC 636.294:575.174:575.113:577.2.08:51-76

doi: 10.15389/agrobiology.2015.6.756eng

Acknowledgements:
The equipment of Bioresources and Bioengineering Center of L.K. Ernst All-Russian Research Institute of Animal Husbandry was used.
Supported by the Russian Science Foundation, project № 14-36-00039

DEVELOPMENT OF MULTIPLEX MICROSATELLITE PANEL TO ASSESS
THE PARENTAGE VERIFICATION IN AND DIFFERENTIATION DEGREE
OF REINDEER POPULATIONS (Rangifer tarandus)

V.R. Kharzinova1, E.A. Gladyr’1, V.I. Fedorov1, 2, T.M. Romanenko3,
L.D. Shimit4, K.A. Layshev5, L.A. Kalashnikova6, N.A. Zinovieva1

1L.K. Ernst All-Russian Research Institute of Animal Husbandry, Federal Agency of Scientific Organizations,
pos. Dubrovitsy, Podolsk Region, Moscow Province, 142132 Russia,
e-mail veronika0784@mail.ru;
2Yakutsk Research Institute of Agriculture, Federal Agency of Scientific Organizations, 23/1, ul. Bestuzheva-Marlynskogo, Yakutsk, Sakha Republic, 677001 Russia,
e-mail vfedorov_09@mail.ru;
3Naryan-Mar Agricultural Experimental Station, Federal Agency of Scientific Organizations,
1a, ul. Rybnikov, Naryan-Mar, Nenets AO, 166004, Russia,
e-mail nmshos@atnet.ru;
4Tuva State University,
36, ul. Lenina, Kyzyl, Tyva Republic, 667000 Russia,
e-mail Shimit62@list.ru;
5North-West Center of Interdisciplinary Researches of Food Maintenance Problems, Federal Agency of Scientific Organizations,
7, sh. Podbel’skogo, St. Petersburg, 196608 Russia,
e-mail layshev@mail.ru;
6All-Russian Research Institute of Animal Breeding,
pos. Lеsnуе Роlyаnу, Рushkin Region, Моsсоw Province, 141212 Russia,
e-mail: lakalashnikova@mail.ru

Received September 18, 2015

Reindeer (Rangifer tarandus), the only member of the genus Rangifer, is one of the most interesting object to investigate genetic diversity. One of the technique of studying the genetic structure of populations and parentage identification is to create panels of STR (short tandem repeats) markers. The aim of the current study was the development of multiplex panel of STR markers and assessment of its application to assign the parents and to study biodiversity of Russian reindeer populations. As a biological material for research we used tissue samples (part of ear’s lobes) of reindeer of Even (EVN, n = 44), Evenk (EVK, n = 44), Nenets (n = 45) breeds and Tyva population (TUV, n = 35). DNA extraction was performed using Nexttec columns (Germany) according to the manufacturer’s instructions. Polymorphism of nine STR markers (NVHRT76, RT9, NVHRT24, RT30, RT1, RT6, RT27, NVHRT21 and RT7) was determined by own procedures using ABI 3130xl DNA analyzer («Applied Biosystems», USA). Statistical analysis was performed in MS Excel 2007 with the plugin GenAIEx v. 6.5, software MSA 4.05, Phylip, v. 3.5c, Treev32 and Structure, v. 2.3.4. The studied populations of reindeer were characterized by relatively high levels of genetic diversity. The average number of alleles per locus was 6.11±0.56 in TUV, 6.67±0.50 in NEN, 8.00±0.76 in EVN and 8.89±0.65 in EVK. The smallest effective number of alleles per locus was detected in TUV (3.37±0.47), the maximal value was in EVK (4.89±0.46 alleles per locus), and EVN and NEN occupied an intermediate position (4.42±0.53 and 3.90±0.38, respectively). The number of alleles in single loci ranged from four in NVHRT21 and NVHRT24 for TUV to twelve in RT7 for EVK and RT1 for EVN. The probability of matching genotypes (PI) for the nine loci ranged from 1.84×10-9 in NEN to 5.9×10-11 in EVK, showing the high power of the proposed marker panel for parentage identification. The calculation of the mean values of similarity coefficient Q in the ith cluster with the most probable number of clusters such as k = 3 and k = 4 (Qi/k) revealed high heterogeneity of genetic structure of studied populations. The highest degree of genetic differentiation was shown for TUV (Q2/3 = 0.899±0.034, Q3/4 = 0.883±0.035) and for NEN (Q3/3 = 0.885±0.031, Q4/4 = 0.813±0.038). The EVN and EVK population were close to each other, and a clear clustering between them was not observed. An estimation of Rst (AMOVA) showed that 11.4 % of the total molecular variability was caused by differences between populations, and 88.6 % was due to individual differences between animals (p < 0.01). Evaluation of degree of genetic differentiation of studied populations, using as criteria the values of Nei’ genetic distances and pairwise comparisons of Fst showed similar trends. TUV population was the most distinct comparing to other populations (DNei = 0,283-0,502, Fst = 0,299-0,452), while it was the most differ from NEN and the closest to EVN. The minimal genetic differences were observed between EVN and EVK (DNei = 0,068, Fst = 0,032). The results show high functional power of the developed STR panel to identify the parentage and to study biodiversity in Russian reindeer populations.

Keywords: reindeer breeds, microsatellites, biodiversity, Rangifer tarandus.

 

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