Population Genetics Of Triplophysa Yarkandensis In The Tarim River,Xinjiang

Triplophysa yarkandensis is an indigenous fish endemic to the Tarim River Basin in the south of Xinjiang.Due to its large size and fast growth,it used to be one of the main economic fishes in Tarim River system.However,in recent decades,because of the reduction of water flow,water salinity aggravating,as well as high-intensity fishing and the invasion of exotic fish caused living space and food competition,the population of T.yarkandensis decreased sharply,and it may become another endangered species following the Aspiorhynchus laticeps and Schizothorax biddulphi.Investigating the population resources by analyzing the genetic diversity,genetic structure and historical dynamics are important for guiding significance in protection and scientific utilization of its germplasm resources.In the present study,using SSR markers,mitochondrial DNA（Cytb,D-loop and COI）markers,as well as SNP markers,we studied the genetic diversity level,population genetic structure,historical dynamic information and phylogeny of of eight natural populations in Qarqan River and Weigan River.The main results were as the following:1.Development of polymorphic SSR markers for T.yarkandensisA total of 10,343 SSR locis were obtained from 46,356 transcript sequences from the gonad transcriptome of T.yarkandensis.105 locis with four-base repeating units,more than five base repeating groups and product size less than 300 bp were screened and used as candidate SSR markers.By primary screening using agarose gel electrophoresis,secondary screening using polyacrylamide gel electrophoresis,and further verification using capillary electrophoresis,ten polymorphic SSR locis were finally obtained.Using these 10 SSR loci,we analyzed eight random samples,a total of 92 alleles were obtained,and the observed heterozygosity（H_o）and expected heterozygosity（H_e）of each locus are 0.312-0.828 and 0.629-0.873,respectively,and the polymorphic information content PIC value are between 0.531 and0.856,indicating that all loci have high polymorphism.Hardy-Weinberg equilibrium test results show that all loci did not deviate from the Hardy-Weinberg equilibrium（P>0.05）,which suggest that the 10 SSR loci can be used for the subsequent population genetics studies.2.Population genetics analysis of T.yarkandensis based on SSR molecular markerBased on 10 newly developed SSR markers,we analyzed the population genetic parameters of T.yarkandensis.The results showed that,the polymorphism information content（PIC）,observed heterozygosity（H_o）and expected heterozygosity（H_e）range were 0.629-0.696,0.607-0.736 and 0.655-0.755,respectively,which suggest a high level of genetic diversity in each population.The genetic distance between populations is between 0.0389 and 0.2090,the genetic differentiation index(F_st)is less than 0.05,AMOVA results showed that,the genetic variations mainly derived from intra-population（97.23%）,which indicate that populations have high genetic homogeneity and an obviously gene flow between populations.The results of population clustering and structure analysis showed that the population from Weigan River and the population from Qarqan River were clustered in different genetic branches,and there were obviously different genetic composition between populations from the two tributaries.The analysis of gene flow among populations showed that the Taitma Lake population（TTM）was the center of dispersal in the middle and lower reaches of the Tarim River,and the dispersal direction was the upper reaches of the Weigan River and the Qarqan River.3.Population genetics analysis of T.yarkandensis based on mitochondrial Cytb,COI and D-loop gene sequencesThe genetic diversity and genetic structure of eight T.yarkandensis populations were analyzed using Cytb,COI and D-loop sequences,respectively.The results showed that 41,51and 22 haplotypes were obtained based on Cytb,D-loop and COI sequences,respectively.The haplotype diversity indices were 0.9247,0.9776 and 0.8829,and the nucleotide diversity indices were 0.0065,0.0115 and 0.0032,respectively,the results suggest that each population have a high genetic diversity.The AMOVA results showed that,the genetic variations mainly derived from intra-population（Cytb,96.57%;D-loop,95.25%;COI,95.12%）,moreover,the genetic distance and the genetic differentiation index F_st value is low.Although there were potential genetic boundaries and genetic differentiation between Weigan River and Qarqan River,the genetic differentiation index was only around 0.05(Cytb,F_ct=0.0540;D-loop,F_ct=0.0471;COI,F_ct=0.0400).The results of population history dynamic analysis showd that,the eight T.yarkandensis populations in Tarim River basin were relatively stable and no obvious population expansion occurred recently.4.Genetic structure analysis of T.yarkandensis population based on SNP markersA total of 119 samples from nine T.yarkandensis populations were genotyped by RAD-seq sequencing technology.Finally,164.81 Gb of data was obtained through the Illumina sequencing platform,and 129,873 candidate SNPs were screened for population genetics analysis.The results showed that a high level of genetic diversity was detected in all populations.AMOVA results showed that the most genetic variations originated from inter-populations(F_st=0.67),and the pairwise F_st values ranged from 0.4579 to 0.8736,indicating that the gene flow among populations was at very low levels.The discriminate analysis of principal components（DAPCs）and Neighbor-Joining（NJ）tree revealed that the nine groups can be divided into two clusters.There is a certain degree of genetic differentiation between the two clusters,but have no obvious clustering phenomenon according to the geographical distribution of the Weigan and Qarqan rivers.The Barrier analysis revealed a genetic boundaries between the two tributaries.The Bayesian skyline plotting indicated that T.yarkandensis populations in Tarim River basin had not experienced genetic bottlenecks,and the effective population size remained stable.To sum up,the genetic diversity level of the T.yarkandensis in Tarim River is relatively high,but there is a certain degree of genetic differentiation and genetic barriers between the Weigan River and the Qarqan River population.This may be due to the impact of habitat fragmentation.Therefore,it is necessary to take effective measures to protect the habitat of the T.yarkandensis to avoid the genetic diversity reducing of the population due to the problem of habitat fragmentation,for example,controlling the development of upstream wasteland,especially controlling the expansion of planting areas of high-water-consuming crops,to prevent the sharp reduction of upstream water supply,and strictly implementing the downstream ecological water replenishment project once a year.