Isolation And Analysis Of Polymorphic Microsatellite Loci In Amur Tiger (Panthera Tigris Altaica)

Amur Tiger(Panthera tigris altaica),the largest living cat in the world, lives in the Russian Far East in the Amur-Ussuri region of Primorski and Khabarovski Krais (States) whilst a few are found in northeast of China and Korea. It is the largest and heaviest subspecies of tiger. With less than 500 Amur tigers left in the wild, this magnificent creature is rapidly going extinct in the wild. Therefore, great conservation efforts are needed to be taken to restore this population both in wild and in captivity. Microsatellites have been proved to be a reliable marker for conservation genetics. In order to better understand of its genetic diversity and kinships, population structure and evolutionary history, as a powerful tool, we developed 20 novel microsatellite loci for Panthera tigris altaica. Genomic DNA was extracted from sixty blood samples collected from the Xiongsen Bear and Tiger Village of Guilin in China.The preliminary results are as follows:1. Based on magnetic beads enrichment protocol, 20 novel tri- and tetranucleotide polymorphic microsatellite loci were isolated from a single individual sample.2. The CDP of the 20 novel loci achieved to 1 .000.The data of every allele from PCR of all loci were dealt with CERVUS. These loci showed a total exclusionary power with both unknown parents (Excl-1) or with only unknown sire (Excl-2) were 0.9934 and 0.9999.3. In the study, we had used the loci to conduct a general survey on the genetic diversity of the 60 captive Amur tigers.A total of 89 distinct alleles were observed. The number of alleles per locus ranged from 2 to 9 with an average of 4.50 and mean polymorphic information content (PIC) was 0.545.The observed and expected heterozygosities of these microsatellites ranged from 0.333 to 0.917 (average = 0.617) and from 0.302 to 0.822 (average=0.609), respectively.4. Tests for Hardy-Weinberg equilibrium at each locus were performed. The loci 421H111 431B12 442A7 442D9 443F2 89B10 and 726H6 showed significant deviation from Hardy-Weinberg equilibrium (P < 0.05). Furthermore, the loci 421B121 431B12 441C6 442A7 89B10 presented lower observed heterozygosity (H_O) than expected (H_E), probably suggestive of existence of inbreeding.5. Through the study, we advanced that the reasons of inbreeding were the unclear pedigree and the unscientific management. In allusion to the present captive tiger status,the study showed that we should investigate genetic diversity of all the captive populations and get the scientific pedigree.With this understanding, together with gene intercommunion across populations,we could adopt logical reproduction management. So we can conserve the genetic diversity of this species better.Besides,establish the individual special gene ID and make a precondition for the management and conservation upon the whole species.