Population Structure Of Dosidicus Gigas In The Southeast Pacific Based On Molecular Markers

Jumbo flying squid,Dosidicus gigas,widely distributed in the Eastern Pacific Ocean,is one of the largest and most abundant species of Ommastrephidae and provides a high-quality protein source for humans.D.gigas suoprts the major comical fisheries of coastal countries including Mexico,Ecuador,Peru,Chile,and the biggest jigging fisheries in the high seas off Eastern Pacific.Understanding the population structure and genetic diversity of the squid population is the basis for the scientific and reasonable assessment and management of the squid fishery resources and the sustainable development and utilization.In this paper,mitochondrial DNA（mt DNA）markers and single nucleotide polymorphism（SNP）markers were used to study and analyze the genetic structure of three phenotypes of D.gigas.The main results are as follows:（1）Primers were designed according to the mitochondrial NADH dehydrogenase subunit 2（ND2）gene sequence,and PCR amplified to genetically analyse the ND2gene sequences of 90 mature D.gigas from three phenotypic groups:large,medium and small.A total of 33 haplotypes were detected in 90 ND2 gene sequences.A total of 33 haplotypes were detected in the ND2 gene sequence,and the sequencing results showed that the haplotype diversity index（H_d）of the three phenotypic groups ranged from 0.768-0.872 and the nucleotide diversity index（P_i）ranged from0.00186-0.00285,showing a high haplotype diversity index and a low nucleotide diversity index.The genetic variation observed within populations reached 100.45%.,The value of genetic differentiation coefficient F_st was less than 0.05,P values were higher than 0.05,and the gene flow（N_m）was far more than 1,indicating that there is no significant genetic differentiation among the different phenotypes of D.gigas in the Southeast Pacific,and there may be frequent gene exchange between populations.The neutrality tests（Tajima’s D and Fu’s F_s）were both highly significant negative and every nucleotide mismatch distribution curves showed a significant single peak,speculating that the D.gigas may have historically experienced rapid population expansion events.（2）Using GBS（Genotyping-by-Sequencing）simplified genomic technology to develop SNP markers for 90 individuals of matured D.gigas with three phenotypes of large,medium and small in the Southeast Pacific.A total of 392 G of raw data were obtained based on Illumina paired-end sequencing,with an average of 4.36 G per sample and a base quality Q30 ratio of 92.45%.After screening,41914 high-quality SNP markers were finally obtained.The observed heterozygosity（H_o）and expected heterozygosity（H_e）of large,medium,and small phenotypes based on SNP markers are 0.21210 and 0.26039,0.21518 and 0.26826,0.21512 and 0.26673,respectively,and the observed heterozygosity is low,suggesting low genetic diversity in the D.gigas.Principal component analysis（PCA）showed no significant clustering between the different groups of D.gigas,with three groups of samples are mainly concentrating in one area,The value of genetic differentiation index F_st was less than0.05,At the whole genome level,it was indicated that there was no significant genetic differentiation among the large,medium and small phenotypes of D.gigas in the Southeast Pacific.The optimal group number was determined to be 2 according to the K value corresponding to the lowest point of CV error（Cross Validation Error）,which reflected that the samples of the squid were probably from two primitive ancestors.The results of the study on effective population size changes in the population history show that the southeast Pacific D.gigas population may have experienced rapid population expansion between 20000-6000 years ago,the calculation results of the differentiation time show that the medium phenotype group and the large phenotype group D.gigas diverged from the population 800,000 and 600,000 years ago,respectively.Based on the characteristics of historical environmental changes during the Last glacial period,it is speculated that the population expansion and differentiation of D.gigas are related to dramatic historical climate changes（3）The SNP markers of 58 D.gigas individual developed based on GBS technology were further screened and verified by mass spectrometry.A total of 101SNP makers out of 107 candidate SNP markers were detected in the sample,and all of them were found to be polymorphic and showed bi-allelic polymorphism.H_o and H_eranged from 0.0000 to 1.0000 and 0.034 to 0.5000,respectively.The PIC ranged from0.0333 to 0.3750.Twenty-five loci significantly deviated from the HWE after a Bonferroni correction（p<0.001）.These novel SNP markers can be obtained by PCR amplification followed by a rapid test,and would be useful for further population genetic analyses and understanding of this important species,which will enable the sustainable utilization of D.gigas resources.