| The Dybowski's frog(Rana dybowskii)is an amphibian which is mainly distributed in Changbai Mountains in Northeast China.In recent years,the demand for the Dybowski's frog is rising year by year,so the wild ones are becoming more and more precious.Artificial breeding of Dybowski's frog has been widely carried out in Liaoning province,in order to prevent wild resources from being influenced during introduction and farming,it's important to study on the genetic diversity,genetic structure and the difference between wild frog and artificial-bred ones,and molecular markers are ideal for doing this research.Microsatellite DNA in nucleus and cytochrome oxidase I(COI)gene in mitochondria were used respectively to investigate the genetic diversity of Dybowski's frog populations,which were sampled in 4 regions,respectively Fushun,Benxi,Tieling and Dandong in Liao-ning.(1)8 polymorphic loci were selected since they could be stably amplified,and average polymorphism information content(PIC)of the loci was 0.6127.Locus Rpi10O was mono-morphic in Fushun,Benxi and Tieling population,except for Dandong population.87 alleles were observed in 4 populations,and the average was 8.89.The average effective allele(NA),average observed heterozygosity(HO),average expected heterozygosity(HE)were 4.5989,0.3078 and 0.6480,respectively.HE in Dandong population was the highest among all the populations,while in Fushun population it was the lowest.Some unique alleles were also ob-served in the populations.Only locus Rpi104 was in Hardy-Weinberg equilibrium(P>0.05),the rest ones significantly deviated from Hardy-Weinberg equilibrium in the populations(P<0.01).(2)Sequences that were 630bp in length were obtained for analysis.The proportion of base T,C,A,G were 26.9%,30.4%,24.9%and 17.8%,respectively,transition/transversion(si/sv)was 12.8,the number of polymorphic sites was 95,the number of parsimony informa-tive sites was 13.Number of haplotypes was 24,nucleotide diversities(?)were ranged from 0.414±0.124 to 0.846±0.041 in the populations.Tajima's Ds were ranged from-1.4676 to-1.0521,which were not significant except in Dandong population,and this is conformed to neutral mutation.(3)Significant genetic differentiations were observed among populations due to the analysis based on SSR data(FST>0.25).Phylogenetic Neighbour-Joining tree based on the pairwise Nei's standard distances were used to verify the results.There were differences be-tween results from SSR data and COI data,the same result is Benxi population had greater genetic distance to other populations.Results of AMOVA from SSR data and COI data shows that 71.85%and 84.33%of the total genetic variation can be explained by differences within populations.(4)The genetic diversity of Tieling population(farmed)was compared with other popu-lations(wild),and the results showed the genetic diversity of Tieling population is similar to that of other groups.So it is reasonable that Dybowski's frog be farmed in this artificial method.This study investigated the genetic diversities and differentials of the wild and artifi-cial-bred Dybowski's frog respectively,provided theoretical evidence for genetic diversity protection and farming methods. |
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