Quantitative Trait Loci Identification For Growth Habit-related Traits Using The Linkage And Genome-wide Association Analysis In Peanut (Arachis Hypogaea L.)

Peanut,known as Arachis hypogaea,is a grain legume crop that originated from South America and is now grown around the world.Plant growth habit is an important and complex agronomic trait and is associated with yield,disease resistance,and mechanized harvesting in peanuts.There are at least two distinct growth habits?erect and spreading?and several intermediate forms existing in the peanut germplasm.To understand the genetic architecture of the peanut growth habit,it was decomposed into five related traits,including lateral branch angle?LBA?,main stem height?MSH?,lateral branch length?LBL?,extent radius?ER?,and the index of plant type?IOPT?.Because peanut growth habit related-traits are the quantitative traits,and hence there is always an interaction between the genes underlying these traits and environment.This slows the gain in research on the genetic mechanism of peanut growth habit related-traits by traditional molecular genetics.However,with the rapid development of high-throughput sequencing technology and the reduction of sequencing cost,more and more species including peanut have had their entire genomes sequenced.Hence,the combination of high-throughput sequencing technology and traditional mapping methods has become an efficient method for breeders to map QTLs and mine genes related traits.In this study,a segregating F₂ population was developed from a cross of“Jihua 5”and“M130”,35 prostrate plants and 35 erect plants were selected to construct two DNA pools.The technology of a specific length amplified fragment sequencing was used to sequence the specific DNA fragments in the two DNA pools.After which molecular markers associated with peanut growth habit were obtained by screening the mutation sites of the specific DNA fragment.Subsequently,a recombinant inbred line population?F₈?was developed and a new high-density genetic linkage map was constructed by using specific locus amplified fragment sequencing technology.Quantitative trait locus?QTL?analysis of the growth habit-related traits was conducted based on phenotyping data from seven environments.Finally,a natural population of 103 U.S.mini core collection were selected as test materials,and the Affymetrix2.0 chip was used to screen polymorphic SNPs and analyze population structure,genetic relationship and linkage disequilibrium.Based on the optimization model,five peanut growth habit-related traits?LBA,MSH,LBL,ER and IOPT?were analyzed by GWAS at two environments to dentify SNPs related to traits,speculate candidate genes related to peanut growth habit,and analyze the genetic basis of peanut growth habit.The main purpose of this study is to mine the QTLs associated with peanut growth habit-related traits and provide theoretical basis and reference for molecular marker-assisted breeding of peanut growth habit breeding.The main results are as follows:1.Using the BSA-seq method,a total of 1911 candidate polymorphism SLAF labels were obtained by screening between the prostrate pool and the erect pool.Among these were 1023high-quality candidate polymorphism SLAF labels in genome A.Chromosome A09 had the largest number?515?with chromosome A01 having the smallest?18?.Meanwhile,888high-quality candidate polymorphism SLAF labels in genome B.Chromosome B04 had the largest number?192?with chromosome B07 having the smallest?21?.The candidate polymorphism SLAF labels were analyzed by SNP_index method.The correlation threshold after fitting the prostrate pool and the erect pool was 0.5823,and three SLAF labels significantly related to prostrate-erect traits were obtained.The polymorphic SLAF label associated with the prostrate-upright trait was located on chromosome Araip.B05 and the genome region was 142,610,834-146,688,220 with a size of 4.08M.2.A new high-density genetic linkage map containing 2808 single-nucleotide polymorphism markers distributed on 20 linkage groups with a total length of 1308.20 cM and an average inter-marker distance of 0.47 cM was developed.The quantitative trait locus?QTL?analysis of the growth habit-related traits was conducted based on phenotyping data from seven environments.A total of 39 QTLs for growth habit-related traits was detected on 10chromosomes explaining 4.55%²7.74%of the phenotypic variance,in which 6 QTLs were for lateral branch angle,8 QTLs were for extent radius,7 QTLs were for the index of plant type,11QTLs were for main stem height,and 7 QTLs were for lateral branch length.Among these QTLs,12 were co-localized on chromosome B05 spanning an approximately 0.17 Mb physical interval in comparison with the allotetraploid reference genome of“Tifrunner”.Analysis of the co-localized genome region has shown that the putative genes are involved in light and hormones and will facilitate peanut growth habit molecular breeding and study of peanut domestication.3.One hundred and three?103?U.S.mini core collection were genotyped using an Affymetrix array.After filtering out SNPs with low quality?call rate<0.95?and SNPs with MAF<0.05,a total of 12,342 SNPs were obtained.The SNP density of chromosome B09 was the highest?0.10 M/SNP?with 1,428 being the number of SNPs.On the other hand,the density of SNPs of chromosome A10 was the lowest?0.37 M/SNP?with a SNP number of 293.The polymorphic information content?PIC?value ranged between 0.26 to 0.30 with the average being 0.28.Population structure,phylogenetic relationship and principal component analysis of 12342selected SNPs were performed.The population structure analysis showed that the peanut mini-core collection has four subgroups,G1,G2,G3 and G4.When r² was equal to 0.2,LD decay distance was 0.16M.A total of 91 associated SNPs were detected by GWAS in two environments.They were distributed on chromosomes A01,A02,A03,A04,A05,A06,A07,A09,A10,B04,B05,B06,B07,B08 and B10.The 597 candidate genes were detected which play important roles in biological processes,hormone signaling,growth and development.4.The SNP?AX-147251085,144,353,467?associated with LBA by GWAS method was overlapped with the results in BSA-seq through combined analysis of GWAS and BSA-seq.Based on LD decay distance,the genome range related to LBA on B05 was shortened to144,193,467-144,513,467.Three candidate genes related to F-box family proteins?Araip.E64SW,Araip.YG1LK and Araip.JJ6RA?and one candidate gene related to PPP family proteins?Araip.YU281?are involved in plant growth and development in this genome region.In conclusion,BSA-seq was used to analyze the association with LBA,the high-density genetic map of peanut was constructed and QTL mapping was conducted for phenotypic identification of peanut growth habit-related traits in 7 environments,and peanut Affymetrix 2.0chip was firstly used for GWAS analysis of peanut growth habit-related traits.A large number of SNPs associated with peanut growth habit-related traits were identified.These findings will provide theoretical basis for dissecting the genetic basis of peanut growth habit,and genetic improvement of peanut growth,and breeding of new varieties.