Identifying Nitrogen-efficient Chinese Cabbage Varieties And Genome-wide Association Analysis

In the process of Chinese cabbage production,if the blind pursuit of high yield and excessive application of nitrogen fertilizer will lead to a large amount of nitrogen fertilizer loss and serious environmental pollution.Therefore,finding nitrogen-efficient varieties and cloning genes that control nitrogen use efficiency are urgent problems to be solved in future research work.Chinese cabbage nitrogen use efficiency is a complex trait controlled by multiple genes and the environment.This experiment is divided into two steps:Identify Nitrogen-efficient Chinese Cabbage Varietiessixty-eight representative Chinese cabbage cultivars from the main producing areas(Shandong,Beijing,Tianjin etc.)were planted in the field with low(Pure N54 kg/ha)and normal(Pure N 270 kg/ha)nitrogen inputs,respectively in the study.A total of 15 agronomic traits,including plant width,plant height,total weight,outer leaves number,net weight,leafy head height,leafy head width,head leaves number,maximum leaf length,maximum leaves width,petiole length,petiole width,petiole thickness,central axis length and central axis width,and three nitrogen related traits including nitrogen content,nitrogen agronomic use efficiency(NUE)and nitrogen responsivity were calculated and analyzed at leafy head maturation stage.Finally,the relative values of each trait in two nitrogen levels were calculated,and the classification for NUE and nitrogen responsivity classes of the 68 Chinese cabbage cultivars were conducted according to yield,relative yield,and nitrogen responsivity.The results showed that 13 agronomic traits were significantly different between the two nitrogen levels,except for outer leaves number and central axis length.Nitrogen content and NUE also showed significant differences between the two nitrogen levels.There were significant genotypic variations for all the agronomic traits under both nitrogen input conditions,of which,central column length had the largest coefficient of variation(CV)among cultivars,followed by weight-related traits.Nitrogen content,nitrogen agronomic efficiency and nitrogen responsivity also showed significant genotypic variations,of which,CV of nitrogen responsivity was the largest.The means of relative net weight and relative gross weight were the largest,while those of relative outer leaves number,relative plant width,and relative central column length were much smaller.In addition,the 68 tested Chinese cabbage cultivars were categorized into four NUE classes(Nitrogen efficient under the two nitrogen conditions,NET;Nitrogen efficient under the low nitrogen condition,NEL;Nitrogen efficient under the normal nitrogen condition,NEN;Nitrogen inefficient under the two nitrogen conditions,NIT)according to the yield under both nitrogen supply conditions,and four nitrogen sensitive types(Nitrogen insensitive type,NIS;Nitrogen low-sensitive type,NLS;Nitrogen moderate-sensitive type,NMS;and Nitrogen high-sensitive type,NHS)according to relative yield and nitrogen responsivity.Using both NUE and nitrogen sensitivity classification criteria mentioned in the study,the nitrogen response characteristics of different Chinese cabbage varieties can be more clearly evaluated.The study will provide useful information for further studying the molecular mechanisms regulating NUE in Chinese cabbage andbreeding new Chinese cabbage cultivars with high NUE.Genome-wide association analysis of nitrogen efficiency-related traitsThe study will provide useful information for further studying the molecular mechanisms regulating NUE in Chinese cabbage and breeding new Chinese cabbage cultivars with high NUE..Therefore,based on genotyping by sequencing data from 149 B.rapa inbred lines with a high purity and broad genetic background,many single nucleotide polymorphisms(SNPs)and insertion-deletion(Indels)were selected out for further study.A genome-wide association study(GWAS)was then performed in order to get the candidate genes and to provide the basis for molecular breeding ? Genome-wide association analysis showed that356,077 SNPs and 193,963 InDels were obtained from the 10 chromosomes of Chinese cabbage.Among them,SNP and InDel detected on chromosome A09 were the most,which were 47,345 and 29,017 respectively.The SNP and InDel detected on A10 were the least,18,034 and 11,958 respectively.The average frequency of variation of SNP and was 1/722 bp and the average frequency of variation of InDel was 1/1326 bp,The results of genome-wide association analysis showed that 356,077 SNPs and 193,963 InDel were obtained from 10 Chinese cabbage chromosomes.Among them,A09 chromosomes detected the most SNPand InDel,with 47,345 and 29,017 respectively;A10 detected the least SNP and InDel,with 18,034 and 11,958,respectively.The average mutation frequency of SNP is 1 / 722 bp,and the average mutation frequency of InDel is 1 / 1326 bp.The results of Manhattan and QQ plots show that a total of 95834 SNP loci(P ? 0.05)associated with the important traits of Chinese cabbage were detected in 2018 and2019.Under normal nitrogen supply conditions,a total of 29310 sites were detected in 2018,and 29800 sites were detected in 2019,and the number of sites for each trait varied significantly,ranging from 29 to 10280 sites,which can be repeated within two years The total number of detected loci is 14686,the number of loci for various traits ranges from 0 to 6323,and the contribution rate is from10% to 88%;under low nitrogen conditions,7674 loci were detected in 2018 and29050 in 2019 For each locus,the number of loci for each trait varies significantly,ranging from 10 to 5502 loci.The total number of loci that can be repeatedly detected within two years is 1581,and the number of loci for each trait varies from0 to 867.The contribution rate ranges from 10% to 88%.Among them,42 sites of nitrogen responsiveness correspond to 11 genes,and genes are annotated.