| The clam Meretrix petechialis is a commercially important marine bivalve mollusk,but its population has been greatly reduced due to over-exploitation and habitat destruction.The genetic protection of M.petechialis is essential for the sustainable development of natural resources.In recent years,with the deepening of research,biological genetic diversity is of great significance for the protection,development and utilization of species.In this experiment,the genetic background of M.petechialis will be studied through the following aspects,and the genetic differences between different geographical groups and different shell colors of M.petechialis will be compared to provide a scientific theoretical basis for the research and breeding of M.petechialis germplasm resources.In this study,a set of novel EST-derived microsatellite makers were developed and characterized in the clam M.petechialis.The number of alleles at each locus ranged from 4 to 30with an average of 10.9 alleles per locus.The observed and expected heterozygosities varied from 0.100 to 1.000 and from 0.538 to 0.973,with an average of 0.713 and 0.750,respectively.The PIC value ranged from 0.423?Mmt09?to 0.956?Mmt47?,with an average of 0.695.The cross-species amplification transferability of nine microsatellite loci to three closely related species ranged from 29.17%to 100%.These microsatellite loci will be useful for further investigation of population structure and conversation genetics of this species.In the current study,we conducted a microsatellite analysis study on nine populations of M.petechialis and six novel EST-derived microsatellite markers were selected for assessment.Based on the experimental results,the population differentiation degree was analyzed.And research and evaluate its genetic diversity.We have detected a total of 165 alleles.At each of these 6 loci,the minimum number of alleles is 4 and the maximum is 30 for each locus.The observed mean minimum is 0.717,the maximum is 0.861,and the average of the expected heterozygosity is 0.797,the maximum is 0.856.According to overall FST=0.214,P<0.01,it can be concluded that genetic differentiation is significant.The analysis methods used in the analysis and research in this paper have the average of the formula,the unweighted pair method?Neighbor-joining?.The result shows that M.petechialis populations from GX are the farthest populations,indicating a certain degree of genetic variation among individuals in each population.This study will provide a basis for the domestication and cultivation of genetic diversity of M.petechialis population in China,also provide the cultivation of clam germplasm resources in protected.To provide valuable insights into management and conservation of M.petechialis,we investigated the genetic variation and population structure of M.petechialis by analyzing samples from nine geographical populations.In this study,the genetic diversity and differentiation of nine populations of M.petechialis was assessed using the mitochondrial cytochrome oxidase subunit I?mtCOI?gene.A total of 90 COI sequences were obtained and each COI sequence was 699 bp in length.Fifty-one haplotypes were identified with 10haplotypes shared among populations.The haplotype diversity was highest in FJ,PJ,and JS?0.9778±0.0540?and lowest in DD?0.7778±0.1374?.The nucleotide diversity was highest in PJ?0.4534±0.2405?and lowest in JS?0.0062±0.0041?.Neutral test?Fu's Fs?and mismatch distribution analysis revealed that the hard clam experienced a population expansion event.Analysis of molecular variance?AMOVA?indicated that 91.7%of the genetic variance was within populations and 0.52%of the variance was among populations,demonstrating significant genetic differentiation among populations?P<0.05?.The Neighbor-joining tree showed that the haplotypes were not clustered according to geographical location,but some haplotypes from the same or neighboring locations grouped together.The results obtained in this study provide useful information on the genetic diversity and population structure of M.petechialis and shed light on the management and protection of resources M.petechialis in the northwestern Pacific.The genetic diversity of 9 populations of M.petechialis was analyzed by SRAP?sequence-related amplified polymorphism?molecular markers.Eight pairs of primer combinations with clear and polymorphic bands were screened out from 105 pairs of primer combinations.The number of sites detected by each pair of primer combinations was 12-23,and132 positions were detected in 9 M.petechialis populations.The JS population had the highest percentage of polymorphic loci?27.27%?.The Nei'S gene diversity of the nine M.petechialis populations ranged from 0.0647 to 0.0793,with the SD population being the lowest and the NK population being the highest.The Shannon'S information index is between 0.1023 and 0.1202,with the lowest SD population and the highest JS population.The unbiased genetic distance of Nei'S between 9 populations was 0.0243-0.0570,the genetic similarity was 0.9446-0.9760;the genetic distance between GX and SD population was the largest?0.0570?,the genetic relationship was far,and the genetic distance between SD and JS population was the smallest?0.0243?,the kinship is closer.This provides a scientific basis for M.petechialis germplasm conservation and improved breeding.In order to screen the molecular markers associated with the shell color,gene expression of two shell colors of M.petechialis were analyzed by SRAP-cDNA.The two shell color genomes of M.petechialis were amplified using 30 primers'combinations,and 11 pairs of primers showed differential fragments,and by recovery,cloning and sequencing,18 different differential sequences were obtained.The sequencing results were analyzed by BlastX.It was found that Ton-dependent receptor protein may be involved in shell color regulation.SCAR primers were designed according to the difference sequence,and PCR amplification was performed in the"yellow"and"red"M.petechialis,and 4 pairs of differential primers were obtained.Using the population to verify the four markers?Me1/Em2,Me2/Em3,Me4/Em11 and Me4/Em12?,it was found that Me1/Em2 and Me2/Em3 were positive in the"yellow"M.petechialis population,whereas Me4/Em11 and Me4/Em12 are positive in the"red"M.petechialis population,and all four markers can be used as M.petechialis shell color related markers.This provides a theoretical basis for studying the different shell color regulation of M.petechialis,thus revealing the molecular mechanism of M.petechialis shell color more comprehensively were analyzed by SRAP-cDNA.The two shell color genomes of M.petechialis were amplified using 30 primers'combinations,and 11 pairs of primers showed differential fragments,and by recovery,cloning and sequencing,18 different differential sequences were obtained.The sequencing results were analyzed by BlastX.It was found that only one fragment shared high homology with memory related protein-2 and no protein and gene related to shell color were found.SCAR primers were designed according to the difference sequence,and PCR amplification was performed in the"yellow"and"red"M.petechialis,and 4 pairs of differential primers were obtained.Using the population to verify the four markers?Me1/Em2,Me2/Em3,Me4/Em11 and Me4/Em12?,it was found that Me1/Em2 and Me2/Em3 were positive in the"yellow"M.petechialis population,whereas Me4/Em11 and Me4/Em12 are positive in the"red"M.petechialis population,and all four markers can be used as M.petechialis shell color related markers.This provides a theoretical basis for studying the different shell color regulation of M.petechialis,thus revealing the molecular mechanism of M.petechialis shell color more comprehensively. |
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