| As one of emerging animal models, Tree Shrews have been applied widely for biomedical research. These animals are phylogenetically close to primates in evolution. Due to some characteristics were exhibited from the Tree Shrew, such as small size, easy to domestication, good reproduction ability, spontaneous diseases and experimental animal for HBV, HCV infection. Tree Shrews have been applied for lots of studies including virology, neurology, immunology, oncology and social biology and so on. However, people showed more interest to them in the applied research than the animal itself. Especially, there were rare studies about their genetic background, so that Tree Shrews were limited to perform standardization and application as experimental animal. Here, we used respectively RAPD (Random amplified polymorphic DNA) and microsatellite DNA markers to analyze genetic structure of the Tree Shrews (Tupaia Belangeri Chinensis) population, and estimated its genetic diversity. The two molecular makers could provide the theoretical basis and genetic detective methods for us to promote the standardization of laboratory animal of Tree Shrews.For the lack of Tree Shrew genetic information and unknown genome, we had to establish the RAPD genetic markers to understand the genetic diversity of the Tree Shrews population.40 random oligonucleotide primers were first optimized. Then the primers, with effectively analyzing the Tree Shrews RAPD loci, were used for PCR amplification. The results were analyzed with Popgene 1.32 and RAPDistance Package Version 1.04 software for the population genetic characteristics of the Tree Shrews. A total number of 113 RAPD loci in 20 perfect RAPD primers were detected, and 69 of 113 sites were polymorphic, amounting for 61.1%. The average genetic similarity index and the average genetic distance were 0.8307 and 0.1693 respectively, individual genetic distance between 0.09-0.27. Genetic diversity was 0.1864 (male) and 0.1470 (female) respectively, with an average of 0.1667. Partition of genetic variation indicated that 48.29% was distributed within population, and 51.71% between male and female populations. Cluster analysis by NJ tree showed that T15,T33 and T47 individuals were classified into one large group, another gathered by the rest. At the same time, there was intercross among male and female individuals. The results also indicated that the Tree Shrews population had higher genetic diversity, and the selected 20 primers of RAPD were available to analyze the genetic diversity and monitor the genetic variation of the Tree Shrews population.With the advantages of the microsatellite DNA maker, such as high polymorphism, codominant inheritance, and PCR amplification easily and steadily, we established the microsatellite genetic maker method to further analyze the genetic polymorphism of Tree Shrews population using a few isolated microsatellite loci from other species of Tree Shrews from papers. Moreover, we could verify whether the results of two genetic test methods between RAPD and microsatellite DNA was consistent. Selected 9 from 11 microsatellite loci could amplify clearly and steadily PCR products for 60 individuals of Tree Shrews by 12% non-denaturing (neutral) polyacrylamide gel electrophoresis test. Then the genetic index for 9 microsatellite loci, such as the number of alleles, allele frequency, polymorphism information content (PIC), heterozygosity were calculated.51 alleles were detected from the Tree Shrews population in 9 microsatellite loci. The average number of alleles was 5.667. The expected heterozygosity of TG22 was the highest 0.8936, the lowest for the TG1 of 0.5042, the average of 0.7648. The polymorphism information content (PIC) of 9 microsatellite loci ranged from 0.4985 to 0.8896, with a mean of 0.7588. It showed that the Tree Shrews population had higher genetic diversity. Besides, the amplified PCR products of 6 selected microsatellite loci were sequenced. After sequencing, we could find that the motif of microsatellite loci were mainly made of the perfect and imperfect (AC)n or (AC)n short tandem repeat. The sequence homogeneity varied from 75% to 93%. The results indicated that the selected 9 microsatellite loci with high genetic polymorphism could be used to analyze the genetic diversity of the Tree Shrews population.These results showed that the Tree Shrews population had higher genetic polymorphism. Here, two genetic testing methods including RAPD and microsatellite DNA genetic makers had been established respectively, which could be applied to analyze the genetic structure of the Tree Shrews population and estimate their genetic diversity. The Tree Shrews with a high polymorphism facilitate to the propagation of animal breeding, but if we want to use widely the Tree Shrews for biomedical research, we must promote the process of standardization of laboratory animals to ensure consistency and reliability of experimental results.
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