Mapping Of Quantitative Trait Loci For Aspergillus Flavus Resistance In Peanut

Peanut(Arachis hypogaea)grown throughout the globe for its protein and oil contents.It is nature’s zero waste crop;each part of the plant is used for various purposes.Peanut kernels contain about 48 to 50 % oil and 25 to28 % protein,providing a rich source of energy for a large group of the human community throughout the world.Its kernels are consumed as raw,boiled or roasted,and also in the form of culinary oil.Being a rich source of human diet(antioxidants,minerals and vitamins),animal feed(oil pressings,green straw and pods),industrial raw material(oil cakes and fertilizer),and soil fertility(atmospheric nitrogen fixation),peanut is a brilliant cash crop for both domestic markets as well as foreign trade.Having inimitable importance in food and feed security,the production and quality of peanut are greatly threatened by aflatoxins(AFs)contamination.AFs are carcinogenic compounds produced mainly by Aspergillus flavus and Aspergillus parasiticus.AFs causing several lethal mutations in both humans and animals,which can ultimately result in liver cancer,retarded growth,immune suppression,and speed up in the progression of acquired immune deficiency syndrome(AIDS).AFs exposure may add to high viral loads,badly affecting the normal liver functioning in HIV-positive patient and thus promoting disease progression.No magic bullet found yet to solve this problem,however,several techniques have been tested to minimize and control AFs contamination including different physical,chemical,and biological preventions.Many biological control agents,including nontoxigenic fungal strains,yeasts,and bacteria have been applied and considerable achievements gained,but for the complete eradication of this intricate problem,stable resistant cultivars development is the best option.Unfortunately,in case of cultivated peanut development of AFs resistant cultivars remained a bottleneck,due to the dearth in availability of consistent resistance resources.The complexity of this problem is further intensified by the interference of various other factors with the AFs production pathway.Mapping resistant QTLs being one of the strategies to develop resistant cultivars against A.flavus infection and AFs contamination,here we map two resistant QTLs.For closely locating the QTL of interest the key is to get stable mean phenotypic data against the trait of interest and construct a high-density genetic map(HDGM).Our goal was to identify host plant resistance to the fungal infection and AFs contamination in order to mitigate this food safety issue.Keeping in mind the complexity of the trait we characterized a large number of recombinant inbred lines(RILs),obtained from two highly contrasting parents against A.flavus infection,for multiple seasons.These two RILs mapping populations,named R,containing 340 RILs and XY,consist of 900 RILs,were developed through single seed decent(SSD)method via crossing the same parents,Xinhuixiaoli(XHXL)and Yueyou 92(YY92),but different times of crossing.The male parent XHXL is highly resistant to A.flavus infection,a landrace belonging to A.h.ssp.fastigiata var.fastigiata,collected at Guangdong province of China.It was identified by us showing stable resistance to A.flavus both in pre-harvest fields and post-harvest inoculation for many years.The female parent YY92,A.h.ssp.fastigiata var.vulgaris,highly susceptible to A.flavus infection,is a Chinese cultivar developed by Guangdong Academy of Agricultural Sciences.Furthermore,the phenotyping step was divided into two portions.In one portion the peanut kernels were in vitro inoculated and characterized under control environment in the laboratory,while in the second portion inoculation was carried out in the field at flowering stage of the crop and then through visual observation the resistance and susceptibility level were recorded without further inoculation.Based on infection index 210 RILs were found consistently within resistance range under multiple seasons.Using specific-locus amplified fragment sequencing(SLAF-seq)technology,a total of 1,434 Million reads of raw data were generated within the range of314 to 414 bp with Q30 values of 86.64 % and 43.62 % GC content.The total number of high-quality SLAFs found were 790,026,of which 56,011 were polymorphic.Using 19,064 high-quality polymorphic SLAFs an HDGM was constructed for cultivated peanut,consisted of 5,022 single nucleotide polymorphism(SNP)markers covering all 20 linkage groups spanning 2,231.25 c M with an average adjacent marker distance of 0.44 c M.Using the mean phenotypic data and the genotypic data generated via SLAF-seq,two resistant QTLs,q RAF-3-1 and q RAF-14-1,were mapped on linkage groups(LGs)A03 and B04,respectively.QTL q RAF-3-1 was mapped within 0.38 c M with more than 19 % phenotypic variance explained(PVE %).In comparison with the reference genome,these two QTLs,q RAF-3-1 and q RAF-14-1,were located within a physical distance of 1.45 Mega base pair(Mbp)and 2.22 Mbp,harboring 67 and 137 genes,respectively.Genes identified in the q RAF-3-1 region included those coding for WRKY transcription factor,putative disease resistance RPP13-like protein 1,and cytochrome P450 71B34.Using microarray gene expression study,most genes including WRKY transcription factor,cytochrome P450 71B34,pentatricopeptide repeat-containing-like protein,and putative disease resistance RPP13-like protein 1 responded to A.flavus challenge.Several evaluated RILs with significantly low in A.flavus infection identified,which will add to the resistance resource against A.flavus infection and AFs contamination.Our mapped resistant QTLs,its underlying resistant putative candidate genes with potential marker development for marker-assistant selection against A.flavus and AFs contamination will play a significant role in bridging the gap of AFs production in peanut.Moreover,our constructed HDGM will also provide great assistance in fine mapping of other important traits in cultivated peanut.