| Harpadon nehereus belonging to Myctophiformes order,Harpadontidae family,and Harpadon genus,is an economically and ecologically important fish species.In this study,mitochondrial DNA and microsatellite markers were applied to evaluate the population genetic structure and corresponding formation mechanism of H.nehereus collected from China offshore,and provide fundamental genetic data and theoretical for the sustainable utilization of fishery resources of H.nehereus.The main results were listed as follows:1.Characterization of microsatellite loci and polymorphism detection of H.nehereusBased on high-throughput sequencing technology,the transcriptome and whole genome survey dataset were generated and employed to develop microsatellite markers and provide preliminary genomic information of H.nehereus.We applied the transcriptome sequencing technique to identify about 5,652 perfect microsatellite loci from 16.97 Mb Unigenes.Among all these loci,the mononucleotide motif(67.59%)was the most abundant.In transcriptome dataset,a total of 148 repeated units were detected,the copy number of repeat motif was dominated by 10,and the length of microsatellite sequence ranged from 10 bp to 92 bp.The whole genome survey sequencing obtained a total length of 1.36 Gb Scaffolds.The genome size of H.nehereus was estimated to be 1,315 Mb by K-mer analysis.A total of 1,027,651 microsatellite loci were identified using MISA software,in which the most frequent repeat motif was dinucleotide with a frequency of 54.35%.On the whole genome wide level,a total of 1,155 repeated units were detected with the copy number of repeat motif dominated by 10 and 22,and the main length of microsatellite sequence ranged from 10 bp to 92 bp.Among the primer pairs designed for 100 loci for PCR amplification,26 loci were polymorphic.These loci detected 179 alleles among 24 fish samples.The average expected heterozygosity(He),observed heterozygosity(Ho)and polymorphic information content were 0.753±0.166,0.761 ±0.115 and 0.713±0.120,respectively.The results showed that H.nehereus has abundant microsatellite loci and high molecular marker development potential.The developed 26 microsatellite markers can be applied in the population genetic diversity researches of H.nehereus in the future.2.Population genetic study of H.nehereus based on microsatellite markersThe genetic structure of H.nehereus populations was evaluated using 6 microsatellite markers.The allelic richness(Rs),heterozygosity(He,Ho)and polymorphic information content(PIC)of these geographical populations were high,showing a high level of genetic diversity of H.nehereus.Significant genetic differentiation was detected between the East and South China Sea populations(SM,QZ ZJ,BH and SY)and the Yellow Sea populations(LYG and NT)based on Fst values.UPGAM tree indicated the populations of LYG and NT formed a separate clade.Structure and AMOVA analysis showed that H.nehereus populations could be divided into two free mating groups with significant genetic differentiation.Furthermore,there is no geographical isolation pattern and recent genetic bottleneck in H.nehereus populations.3.Population genetic study of H.nehereus based on mitochondrial DNA cytochrome b(Cyt b)sequenceThe population genetic diversity,genetic structure and demographic history of H.nehereus collected from 13 sites in China offshore were investigated base on the length of 1073 bp fragment of mitochondrial DNA cytochrome b(Cyt b)gene.The results showed that there were relatively low haplotype diversity(h)and nucleotide diversity(π).No significant genealogical branches corresponding to sampling locations were revealed both in Neighbor-Joining(NJ)and Minimum Spanning(MS)trees.AMOVA and pairwise Fst values indicated no obvious genetic structure and the slight genetic variation mainly originated within individuals.Both mismatch distribution analysis and neutrality tests showed H.nehereus had undergone population expansion,possibly before the last 30000-70 000 years. |
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