| Objectives:Sika deer is subordinated to artiodactyla, cervidae, Muntiacus. Northeasten sika deer (cervus nippon hortulorum), one of the three survival subspecies in China,has important value of medical use and economy value.It has generally forcused on physiochemiacal identity,feeding management,diseases prevention and cure in cervus nippon hortulorum researches,but the researches about the nuclear genes and chromosomes are much less. The analysis of N-banding,the amplified fragment length polymorphism and the Microsatellite DNA can provide the basic information for genetic polymorphisms research of c. n. hortulorum. And our work provide the basic data for the further research on the genetic variation,important economic character and the gene localization of c. n. hortulorum.Methods:(1) The peripheral blood was obtained while sawing the velvet antler. Collect lymphocytes after 72-hours culture. The chromosomes were dealed with N-banding technique,and the result of N-bands of c. n. hortulorum chromosomes was analysed.(2) The peripheral blood was obtained while sawing the velvet antler. Collect lymphocytes after 72-hours culture. The genome DNA of c. n. hortulorum was extracted from the cells.And The genomic DNAs of the 27 different Cervus Nippon hortulorum were analyzed by amplified fragment length polymorphism (AFLP) with 9 kinds of primer combinations and restriction endonucleases EcoRâ… /Mse.(3) The genetic polymorphism of fifteen microsatellite loci of c. n. hortulorum was detected by polymerase chain reaction ( PCR), 5 % nondenaturation polyacrylamide gel electrophoresis and Sanguinetti silver staining method.Results and conclusions: (1)It is banded in centromeres of autosomes. The obvious N-banding was observed in centromeres of the fifth,thirteenth,sixteenth,twenty-sixth and thirty-second chromosomes. There is a piece of bigger and darker area in the arms of X chromosome. All parts of Y chromosome is coloured deeply and the centromere is much deeper than any parts of the chromosome obviersly. All the results indicated the diversity of the N-banding which shown as the location and the dyeabilityof the band. (2) 11443 bands associated with genetic polymorphism among the total 15169 bands were obtained, and the percentage of polymorphic bands was 78.43%, 1271 polymorphic locus were shown per primer combination. The AFLP data showed that the average genetic similarity was 0.7841(0.6809-0.8648). The 27 samples were classified into four groups with cluster analysis. The results of AFLP and cluster analysis concluded that there was high genetic variation, which associated with orientated artificial breed selection and breeding in the population.(3)The results suggests that two of fifteen, RT1 and NVHRT16, microsatellite loci are high polymorphic , and the other microsatellite loci have no polymorphism. It is the conclusions that the two microsatellite loci are effective DNA markers for analysis of genetic diversity, and could be used to direct artificial breed selection.All the results illustrated that N-banding,AFLP and microsatellite technique were feasible and effective method applied to the analysis of genetic polymorphism, identification of species and genetic relationship. The results laid the foundation for the main quantitative trait locus (QTLs) of Cervus Nippon hortulorum.
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