Identification Of Ceel-MHC Class Ⅱ Genes And Analysis Of Genetic Variation For The Hainan Eld’s Deer

Hainan eld’s deer (Cervus eldi hainanus) is endemic in China, once widely distributed in low altitude sloping fields and shrub forests in the Hainan Island. Its population had decreased seriously due to rampant poaching and habitat destruction. Although in recent decades the population has been rebounding, all the deer in existence are descendants of26individuals kept in the Datian Natural Reserve in1976. Therefore, founder effect resulted in low genetic diversity, which made the deer be vulnerable to various diseases and bleak in adaptive ability.Major Histocompatibility Complex (MHC), which is an important part of vertebrate immune system, presents high polymorphism. Moreover, it is a significant indicator of population genetic structure and adaptive capacity, and is also used for disease risk assessment. This research employed RACE (rapid-amplification of cDNA ends) technology to acquire the entire coding sequences of Hainan eld’s deer’s MHC class Ⅱ genes, of which the quantity was also assessed based on the sequence information from gDNA. After designing gene-specific primers, we studied genetic variation in Bangxi population. Main results were as follows:1) Four complete coding sequences, including two Ceel-DRA, one Ceel-DQA1and one Ceel-DQA2, were isolated from the Hainan eld’s deer. They belonged to three MHC class Ⅱ A loci expressed in different organs (e.g. liver, kidney and spleen), which had complete structure and could be coded into normal protein molecules. The features mentioned above were in accordance with that of classical MHC genes, indicating that these three genes were all classical MHC genes in Hainan eld’s deer. Furthermore, no silent gene or pseudogene was discovered when scanning the gDNA samples.2) Six complete coding sequences from MHC class Ⅱ B loci were identified in the Hainan eld’s deer, including:three Ceel-DRB and three Ceel-DQB. They might be derived from one or two Ceel-DRB loci and two or three Ceel-DQB loci respectively. Due to the limitation of experimental materials, the number of B loci could not be precisely determined. However, after referring to the sequence information of exon and intron and PCR scanning of seven individuals, Ceel-DRB*01and Ceel-DRB*03could certainly be alleles. Since Ceel-DQB*01was derived from an independent locus, it apparently differed from the other two Ceel-DQB sequences. Based on the results of gene structure analysis and tissue expression, all these six sequences should be derived from the classical MHC loci.3) After amplifying A genes using the gene-specific primers and B genes with species-specific primers, the genetic diversity of Bangxi population was investigated. Except for the four A gene sequences and six B gene sequences listed above, no additional sequences were found. This result suggested obvious founder effect in the Bangxi population. Thus, it was necessary to intensify gene flow among extant populations.4) Ceel-DRA gene in Hainan eld’s deer was very conservative. The differences in nucleotides and amino acids between its two alleles were0.9%and0.8%respectively. Moreover, DRA exon2was quite conservative among different deer. The amino acid composition of the four DRA sequences from red deer (Cervus elaphus) was identical with that from Hainan eld’s deer. Therefore, the variation level of Ceel-DRA gene in Hainan eld’s deer required little concern, as the lack of practical variation between Ceel-DRA alleles was probably caused by purifying selection.5) Unlike Ceel-DRA gene, the two Ceel-DRB alleles of Hainan eld’s deer were obviously different; the differences in nucleotides and amino acids were2.9%and6.4%respectively. This might be caused by positive selection. In addition, the Bayes tree showed that the sequences from Hainan eld’s deer were mixed with that from red deer and sika deer (Cervus Nippon), which demonstrated trans-species evolution of DRB genes occurred among different deer.6) Since positive selection, which accumulates advantageous mutations to improve the fitness and adaptive potential of a population, have influenced the apparent differences in Ceel-DRB alleles, it was necessary to pay attention to the genetic variation of Ceel-DRB gene in the Hainan eld’s deer in future population management. It would be better to reproduce heterozygous individuals at Ceel-DRB locus as many as possible in breeding program.