Development And Application Of SNP In Large Yellow Croaker (Larimichthys Crocea) Based On Genome And Transcriptome

Large yellow croaker(Larimichthys crocea)is one of the most important economic marine fish in China.However,genetic analysis of the species for quality improvement and molecular-aided selection has been largely hindered by insufficient molecular markers.Single nucleotide polymorphism(SNP)has been served as valuable molecular markers in genetic linkage construction,quantitative traits location,genome-wide association analysis and molecular-aided selection due to their wide genomic distribution and abundant genetic variations.In this study,the major objects are assessing and optimizing the SNP calling pipelines,identifying abundant SNPs markers from genome and transcriptome and utilizing them for QTL mapping of important growth-related traits by next-generation sequencing(NGS)technologies.The major results are as follows:1.Assessing the performance of widely used tools for reads alignment and SNP calling for RNA-seq and investigating the impact of reference sequence,sequencing quantity and sequencing quality on c SNP discovery using simulated reads and benchmark data.The results illuminated that 1)the alignment tools of BWA,Tophat and SOAP2 exhibited significant different performances in mapping ratio and running time;2)c SNP calling by GATK,CLC and SAMtools resulted into divergent sensitivity,false positive rate and c SNP number with/without post-calling filtering;3)reference choice of genome or transcriptome sequence also significantly influenced the accuracy of identified c SNP;4)according to the trade-off between cost and performance,the analysis of the influence of reads quantity and quality illuminated that RNA-seq data of 2.6 Gb with an average read quality of Q25 represented an ideal data set for c SNP development.2.Building up and improving the reduced representation sequencing(genotyping by sequencing,GBS)of large yellow croaker.Using the GBS technology,71105 high-profile putative SNPs were identified from 400 individuals and analyzed their characteristic.Transition was the most base substitution type,accounting for 66 percent of total SNPs number.3.Based on Illumina Hi Seq 2000 RNA-seq technology,29096 high quality SNPs were detected from 72 pedigrees and parents.After statistic and annotation,74% of the SNPs loci were located in the non-genetic interval,a few in the coding region.Meanwhile,based on Roche 454 RNA-seq,5343 putative c SNPs were discovered in transcriptome derived from various organs and tissues of ten individuals.Of the random selected potential 26 SNPs markers,15 loci were found to be polymorphic and showed bi-allelic.4.Using family-based linkage analysis and association study,candidate SNP markers and genes were identified related to important growth traits of croaker.All the three growth-related traits were detected on croaker LG9,LG12 and LG16.Three QTL regions were significantly associated with TW,containing 86 markers and 41 genes.Four significant regions including 56 markers and 25 genes for TL,four significant regions including 36 markers and 18 genes for TL.The identified potential SNPs and genes provided useful information for future research.