The Correlation Between ASIP And MC1R Gene Polymorphisms And Donkey Coat Color And Their Molecular Evolution In Mammals

Animal coat color is an important basis and main mark for identifying animals and individuals in animal husbandry since domestication.Similarly,as one of the important animal husbandry economic livestock in China,the donkey has different economic value,scientific value and production practice value.The three most common donkey coat colors are grey dun,black(NLP),and tri-pink(3-LP),in which there is a "Light Point"(LP)phenotype(light milky white around the eyes,muzzle,and abdomen)unique to other mammals.Previous studies have shown that donkey hair color is mainly determined by the distribution and proportion of eumelanin and melanin produced by melanocytes,which is mainly controlled by two genesASIP(agouti signaling protein)and MC1R(melanocortin-1 receptor)。In the reports about the change of hair color caused by mutation,the LP character of donkey was introduced,but the basic research of molecular biology was lacking.In order to further explore the ASIP gene and the MC1 R gene in the donkey LP phenotypic changes,we collected 283 samples of Chinese donkeys(grey dun phenotype and various five LP phenotypes types),the ASIP and MC1 R gene coding sequence sequencing,after winning mutations,classification and the statistical analysis of the two genotyping the donkey colors and the correlation function of LP phenotypic change.Then,in order to further study the genetic evolution of the two genes in other species,we further analyzed the molecular evolution and selection pressure of ASIP gene and MC1 R gene in the whole mammal.The main results are as follows:By comparing the sequencing results,we identified one nucleotide mutation in the coding region of ASIP gene and MC1 R gene.ASIP gene is non-synonymous mutation c.349T> c,and MC1 R gene is synonymous mutation c.243C>T.ASIP genotype gradually transforms from T/T,through T/C,to C/C(c.349T>C),in linkage with the decreasing size and number of coat color light points,while MC1 R is extremely conserved and has only one synonymous substitution(c.243C>T)in its coding region.A chi-squared test showed that the variation in donkey coat color light points is significantly associated with the ASIP genotypes(p < 0.001),but not with those for MC1 R.These results suggest that the non-synonymous mutation in ASIP may account for the change in size and number of coat color light points in donkeys,whereas the conserved sequence of MC1 R is not a regulatory factor for the six donkey coat color phenotypes.Through the reconstruction of nucleotide and amino acid tree of ASIP gene and MC1 R gene in mammals,we found that MC1 R gene was more obvious than ASIP gene and in the process of mammal evolution.In the analysis of selective pressure,both genes were subjected to different degrees of positive selective pressure.We found that ASIP gene has positive selective sites in primates,cetaceans,artiodactyls,carnivores and kangaroos.The MC1 R gene has positive selective sites in artiodactyls,carnivores,rodents and primates.On the whole,however,through the collection and analysis of the current mammalian gene sequences,ASIP gene and MC1 R gene are relatively conservative in the evolutionary process of mammals,indicating that the corresponding functional protein is very important in the pigment regulatory pathway.Moreover,ASIP and MC1 R,as candidate genes for hair color,are likely to play an important role in the regulation of other traits in animals,such as animal feeding habits,animal behavior,and animal immune regulation.