Genetic Diversity Accessment And Genome-wide Association Analysis Of Important Agronomic Traits In Mandarin Accessions

Citrus（Citrus L.）is one of the most important economic fruit in the world and the third largest trade agricultural product in the world.Citrus has important nutritional value,medicinal value and ecological value.Mandarin is native to China.In the citrus industry in the world,the status of mandarin is second only to sweet orange.There are many varieties of mandarins in China.It is currently in a period of cultivar renewal and structural adjustment.The cultivation of a beautiful appearance,excellent quality,high yield,and easy cultivation has always been the goal of citrus breeding.There have been many reports based on RAPD,AFLP,SSR and other molecular markers for citrus traits association analysis,genome-wide association studies（GWAS）based on single nucleotide polymorphism（SNP）only a few articles have been reported in recent years.To better understand the genetic architecture of the most important breeding target traits in citrus,a population consisting of 240 germplasm accessions was phenotypically characterized for 10 traits and genotyped using genotyping-by-sequencing（GBS）approach,a total of 112,768 high-quality SNPs covered all 9 chromosomes in 240 mandarin accessions were discovered using GBS.Based on phenotype and SNPs,I studied genetic diversity and population structure.GWAS and identification of candidate genes of ten traits Fruit weight（FW）,Fruit length（FL）,Pericarp thickness（PT）,Fruit surface texture（FST）,Total soluble solids（TSS）,Titratable acidity（TA）,Embryo type（ET）,Anther color（AC）,Segment numbers per fruit（SNPF）,Fruit shape index（FSI）,were subsequently performed.The main results are as follows:Statistical analysis of 18 phenotypic traits of 240 mandarins revealed that the mandarin had a high phenotypic diversity.Mandarin accessions from America have higher average contents of TSS and invert sugar（IS）;the mandarin accessions from Japan have larger FW,fruit diameter（FD）and FL and lower seed numbers perfruit（SNF）.Mandarinaccessions from the Yangtze River Basin were larger fruit size,higher contents of the TSS,reducing sugar（RS）and IS than the Pearl River Basin,butlower TSS/TA.Variance analysis for wild germplasms,landraces and bred varieties showed that bred accessions have larger fruits,smoother peels,fewer seeds,lower contents of TA and higher content of sugar content.Both the cluster analysis and PCA suggested genetic variations were presented in different geographical origins.Small fruit type accession from China should be thought as anunique subgroup,Green Peel Mi Ju is an unique citrus accession.Nine phenotypic traits,including FL,FSI,FST,SNPF,petiole length（PL）,leaflamina width（LLW）,the contents of TSS,TA and RS,were selected out as evaluation indexes by stepwise regression analysis.Using GBS technique,a total of 96.3 Gb of sequences were generated from 240 mandarin germplasms,and each sample produced 401.26 Mb in average.PCA analysis and phylogenetic tree analysis were evaluated by 112,768 SNPs,and the results were largely consistent.GBS technology provides an effective and highfeasible approach to assistant the taxonomic classification of 240 mandarin accessions,the classification results are in accordancewith the conclusion based on morphological method.Meanwhile GBS also is a powerful tool for germplasm identification,and can be applied in the new cultivars identification and intellectual property protection.Based on the MLM（mixed linear model）and the significant level of-log10（P）>6,two significant associations with fruit size traits（FW,FL）.The two SNPs located on chromosomes NW₀06262688.1,NW₀06262274.1 and eight candidate genes were found nearby two SNPs;17 candidate genes were found near the five SNPs related to FST,the five SNPs were distributed on chromosome NW₀06262201.1,NW₀06263303.1;The PT related three SNPs were detected on chromosomes NW₀06262339.1,NW₀06261964.1,meanwhile 17 candidate genes were found.A significant SNP locus associated with TSS was successfully identified via GWAS,and three functional genes were found near the SNP distributed on chromosome NW₀06262688.1;The TA related SNP were detected on chromosome NW₀06262688.1,meanwhile 7 candidate genes were found.Six SNPs detected on chromosome NW₀06263303.1 were significantly associated with ET,meanwhile34 candidate genes were found;Ten SNPs,located on chromosomes NW₀06262339.1,NW₀06262022.1,were found to be related to AC,and 55 candidate genes were found near the ten SNPs.SNPF related a SNP was detected on chromosomeNW₀06262022.1,meanwhile one candidate genes was found;Six SNPs distributed on chromosome NW₀06262274.1 that were significantly associated with the FSI,and a total of 27 candidate genes were found near the six SNPs.