| With the biology,medicine and pharmaceutical development, demands of standardization of the experimental macaca monkeys is increasing, and setting up and providing standardization of the experimental macaca monkeys for clear genetic background will provide a more accurate and more reliable results for study of the life sciences. Laboratory Animal Science is the basic and support conditions of biomedical and whole life sciences research, and its development and extent of application is the important sign of measure of a country or region level of science and technology. However, on the whole, it's imbalance for development of experimental animal work in our country, and it's inconsistent with process and the level of standardization, especially like the large-scale experimental animals such as experimental macaca monkeys, and it's much weak to not only the degree of resource, but own research efforts. Experimental animal genetic quality monitoring is the important method to evaluate the quality of animals, in order to validate the genetic characteristics of animal strains, to check whether the existence of heterozygosity, gene mutation and genetic pollution or not, then to determine whether the found objects are in line with the characteristics of biology of groups or not.Because of its features of highly sensitivity, specificity and polymorphism, as well as simple, fast and cheap methods, microsatellite DNA markers was widely applied for genetic quality monitoring of experimental animals and analysis of the genetic structure of diversity, and it is currently recognized the best molecular genetic markers to assess the genetic diversity of species, at the same time, it's also the important methods of experimental animal genetic quality monitoring. The experimental study is mainly screening 30 highly polymorphic, more alleles and chromosome distribution microsatellite DNA loc(iD1S1594, D1S533, D3S3045, D21S1246, D7S513, D6S493, D6S2419, D4S1645, D5S820, D5S1470, D5S1466, D14S255, D15S644, D10S611, D20S171, D12S372, D2S1333, D12S67, D2S146, D11S2002, D11S1352, D9S934, D17S1290, D17S791, D13S797, D18S536, D18S869, D19S571, D19S559 and D16S403)from 100 microsatellite DNA loci through optimizing the PCR amplification conditions of the reaction system, such as the renaturation temperature of monkey DNA microsatellite loci, Mg2+ concentration.Using these 30 polymorphic microsatellite DNA loci, Genetic structure of half of 28 Macaca rhesus in Beijing is analysed, the number of alleles is more than 7, the highest is 11. Using the Popgene3.2 software, indicators such as observed number of alleles(Na), effective number of alleles(Ne), genetic diversity(H)and Shannon information index(I)is analyzed statistically, and then gene frequency is 0.0357~0.3214; observed number of alleles is 7.0000~11.0000, average is 8.0814; effective number of alleles is 5.1579~8.5217, average is 6.5449; genetic diversity is 0.8061~0.8827, average is 0.8453; Shannon information index is 1.7959~2.1682, average is 1.9606, so these monitoring indicators can much better reflect the characteristics of the genetic structure of population diversity, and can construct DNA fingerprinting of polymorphic microsatellite DNA loci in Rhesus monkey populations.At the same time, using these 30 polymorphic microsatellite DNA loci, Genetic structure of half of 28 Macaca irus in Beijing is analyzed, the number of alleles is more than 5, the highest is 10. Using the Popgene3.2 software, indicators such as observed number of alleles(Na), effective number of alleles(Ne), genetic diversity(H)and Shannon information index(I) is analyzed statistically, and then gene frequency is 0.0357~0.2500; observed number of alleles is5.0000~10.0000, average is 7.1501; effective number of alleles is 4.6118~8.3404, average is 6.4255; genetic diversity is 0.7832~0.8801, average is 0.8402; Shannon information index is 1.5702~2.1592, average is 1.8925, so these monitoring indicators can much better reflect the genetic profile of population diversity, and can explain the group have definite heterozygosity and diversity, and can accord with basic requirements of closed colony of laboratory animals, then can construct DNA fingerprinting of polymorphic microsatellite DNA loci in Macaca irus populations.In order to further learn the differences of Macaca rhesus in an inter-subspecies as well as Macaca rhesus and Macaca irus groups, using the screened 15 highly polymorphic microsatellite DNA loci(D1S1594, D1S533, D3S3045, D7S513, D6S2419, D4S1645, D5S1466, D15S644, D10S611, D12S67, D2S146, D11S1352, D13S797, D18S536 and D18S869 ) , DNA polymorphism of the three groups from each 50 Macaca rhesus(male and female half)in Beijing and Sichuan region, as well as 50 Macaca irus reproduced in Beijing is analysed comparatively. The results discover that all loci in three groups show high polymorphism, and the number of alleles is more than 8, the highest is 11. Compared to the number of alleles between the Macaca irus and two Macaca rhesus groups, 7 loci such as D1S533, D3S3045, D6S2419, D4S1645, D10S611, D12S67 and D18S869 have differences. Using the 7 microsatellite DNA can identify effective Macaca irus and Macaca rhesus groups; D12S67 loci and D18S869 loci have differences in the Number of alleles from Beijing and Sichuan region Macaca rhesus groups, so using the 2 loci also can identify effective two Macaca rhesus groups. There are 13 DNA microsatellite loci which have same number of alleles between two Macaca rhesus groups,which is 86.7% of total loci and the results are higher than DNA microsatellite loci which have same number of alleles from Beijing region and Sichuan region Macaca rhesus groups, and the result is 9 and 8, which is 60% and 53.3% of total loci, so these results can explain that genetic distance between two Macaca Rhesus groups is nearer than genetic distance between two Macaca Rhesus and Macaca irus groups.In a word, using these highly polymorphic microsatellite DNA loci screened, the study better reflect the genetic profile of DNA polymorphism macaca monkeys groups, and then construct the fingerprints of DNA genetic polymorphism macaca monkeys groups, and establish the microsatellite DNA markers of molecular genetic monitoring methods between Macaca rhesus and Macaca irus, as well as discriminant analysis methods of DNA genetic background between Macaca rhesus and Macaca irus, and fill the blank of genetic quality monitoring results in domestic laboratory animals Macaca rhesus and Macaca irus, and it is great significant to establish complete Macaca monkey species genetic background resource library, to analyse and research the gene typing, to paternity testing and population genetic distance, to discriminate the different subspecies of Macaca rhesus and Macaca irus species, to select the best ratio of male and female breeding pair, to instruct the breeding production of Macaca rhesus and Macaca irus, to establish the individual macaca monkeys with clear blood pedigree and clear genetic background. The results also provide the strong theoretical basis for molecular genetic markers applying to experimental animals genetic quality monitoring.
|
PDF Full Text Request