Genetic Dissection Of Nitrogen And Phosphorus Efficiency In Relation To Root System Architecture In Brassica Napus L.

In agricultural systems,nitrogen(N)and phosphorus(P)are the essential minerals that limit plant growth.A considerable amount of fertilizer has been utilized to meet N and P requirements to enhance crop production.As a result,improper practices have led to severe environmental issues,low fertilizer utilization efficiency,and excessive annual energy usage.Therefore,it is critical to breed crop varieties that efficiently use nutrients.Roots are responsible for water absorption,nutrient uptake,and anchoring the plant in the soil,thus substantially affecting crop growth and yield formation.Based on the important role of root architectural traits on N and P efficiency in rapeseed,we used linkage mapping and genome-wide transcriptome analysis to investigate the genetic basis for root and shoot biomass traits under low N and P conditions.The main findings are as follows:1.This study evaluated root and shoot biomass traits of 236 rapeseed recombinant inbred lines(RILs)along with their parents,grown under low N(0.5m M N,control(CK)-15 m M N)and low P(0.01 m M P,CK 1m M P)treatments.Heritability estimates for the studied traits under LN/LP ranged from 0.55-0.74 and 0.43-0.73,respectively.Quantitative trait loci(QTL)mapping identified 63 and 49 loci,explaining 4.2-10.7% and 5.0-9.4% phenotypic variances under LN/ LP,respectively.Through QTL-meta analysis,these loci were integrated into 15 significant QTL clusters.Of these,three major and stable QTL clusters involved 13 QTLs that could be repeatedly detected and explained more than10% phenotypic variances,indicating their critical role in N and P’s cooperative nutrients uptake.Moreover,we further investigated candidate genes within these three major QTL clusters.Several candidate genes among them have been associated with root growth and development and/or nutrient stress tolerance.These key loci and candidate genes lay the foundation for deeper dissection of the NP starvation response mechanisms in B.napus.2.The current study evaluated the natural variation of five root and eight shoot biomass traits under a low N treatment(0.5m M N)in a 327-rapeseed association panel,and genome-wide transcriptome analysis was conducted to identify key single nucleotide polymorphisms(SNPs)and candidate genes associated with target traits.The studied root and shoot biomass traits exhibited significant phenotypic differences,with heritabilities ranging from 0.53-0.66,respectively,and most of the traits showed significant correlation with each other.Using 21,242 SNPs,we found 24 significant SNPs integrated into 14 valid QTL clusters associated with root and shoot biomass traits that explained5.7-21.2% phenotypic variance.Candidate genes were mined within the 300-kb region around the lead SNPs.In addition,we performed RNA sequencing to analyze the differential expression of genes(DEGs)transcripts between high and low nitrogen efficiency and high & low nitrogen stress tolerance lines at two developmental stages,7 days(T1)and 14 days(T2),selected from the association panel.The DEGs were then integrated with association mapping to confirm their roles in root development and nitrogen stress.Totally,89,72,and 36 DEGs were identified as HN1LN1/CK-T1T2-specific,HN2LN2/CK-T1T2-specific,and HN1LN1/CK-T1T2 vs.HN2LN2/CK-T1T2-common.The integration of genome-wide association studies(GWAS),weighted gene co-expression network analyses(WGCNA),and differential expression analysis led to the identification of 33 genes that were associated with root and shoot biomass traits.These potential genes affected root growth and development under low nitrogen stress.Previous studies have found that 13 out of 33 candidate genes regulate root development and nitrogen uptake efficiency.These findings revealed the genetic basis underlying nitrogen stress and provided worthwhile SNPs/genes information for the genetic improvement of nitrogen stress tolerance in rapeseed.3.A Brassica napus association panel was evaluated hydroponically for 13 root-related and shoot biomass traits under a low phosphorous level(0.01 m M P to pinpoint key SNPs and candidate genes associated with the investigated traits.Based on the genotypes investigated,significant differences in phenotypic traits were observed,with heritabilities in the range of 0.47-0.56.GWAS analysis identified15 significant SNP-trait associations,integrated into 11 valid QTL clusters for the studied traits that explained 8.2-24.4%.Gene candidates were uncovered within the 300-kb region surrounding the lead SNPs.Moreover,RNA-sequencing analyses were conducted to examine DEGs between high/low phosphorus efficient and high/low P tolerance index accessions at two-time points(T1,T2)and further confirm their role in root growth and phosphorus stress tolerance.In total,406,263,and 438 DEGs were identified as HP1LP1/CK-T1T2-specific,HP2LP2/CK-T1T2-specific,and HP1LP1/CK-T1T2 vs.HP2LP2/CK-T1T2-common.An integrated approach of GWAS,WGCNA,and differential expression analysis identified 25 candidate genes associated with root and shoot biomass traits.These putative genes were considered to be involved in root growth and development during periods of low phosphorus availability.In previous studies,seven out of 25 candidate genes were found to control root growth and phosphorus utilization efficiency.These results explained the genetic basis of phosphorus stress in rapeseed and provided useful information regarding SNPs and genes related to phosphorus stress.4.Based on the integration of QTL Mapping,GWAS,RNA-seq,and WGCNA Analysis,this study identified ten candidate genes that might be associated with root growth and N/P utilization.Six out of10 genes(BnaA09G0061000 ZS,BnaA09G0061200 ZS,BnaA09G0061600 ZS,BnaA09G0061900 ZS,BnaA09G0068200 ZS,and BnaA09G0105300ZS)were found to control root growth and N/P utilization efficiency.Genes related to root growth and development in low N/P stressed environments may be useful in rapeseed molecular breeding and functional studies.