| Soil salinity is a serious threat to agriculture sustainability worldwide.In recent years,association analysis(AM)has been successfully applied to study crop quantitative trait genetic mechanism and find important agronomic genes.In this study,eight salt-tolerant rapeseed(Brassica napus L.)were used to determine the salt tolerance under different Na Cl concentration,and a set of mature hydroponics system was established to evaluate salt tolerance;the salt tolerance of 138 rapeseed inbred lines were evaluated by hydroponics system;the salt tolerance of 368 rapeseed inbred lines were evaluated and the genome-wide association study of salt-related traits at seedling stage with high density SNPs covering the whole genome.The main contents and results are as follows:1.Salt stress of eight rapeseed inbred lines were studied by Na Cl solution of 0 g/L,9.0 g/L,10.5 g/L,12.0 g/L,13.5 g/L and 15.0 g/L Na Cl solution.The results showed that salt stress could inhibit the shoot fresh weight of rapeseed seedlings,and the inhibition was gradually enhanced with the increase of Na Cl concentration.When the concentration was 13.5 g/L,the degree of inhibition between different materials was the largest.Thus,the concentration 13.5 g/L was used as a suitable salt concentration for evaluating the salt tolerance of different rapeseed materials.2.135 g/L Na Cl solution was used to evaluate the salt tolerance of 138 Brassica napus inbred lines.The results showed that salt stress could significantly inhibited the growth of rapeseed seedlings,and the number of fully leaves decreased from 4.99 to 3.16 after salt stress,while the shoot fresh weight decreased from 11.29 g to 4.66 g after salt stress.At the same time,the salt tolerance of different materials was widely varied,the average salt tolerance index of leaf number was 0.64,the variation range was 0.46-0.98,and the salt tolerance index of shoot fresh was 0.45,the variation range was 0.29-0.87.Finally,138 materials were subjected to clustering analysis of salt tolerance,138 materials were divided into four categories,of which 16 were divided into extreme salt-resistant materials;23 materials were classified as salt-resistant materials,18 materials was divided into salt-sensitive materials,the remaining 81 materials were classified as intermediate materials.3.Comparison of seed germination and seedling growth of rapeseed in northeast saline soil and Wuhan native loess.The results showed that northeast saline soil had no inhibitory effect on the seed germination of rapeseed.However,northeast saline soil had a significant inhibitory effect on the seedling growth of rapeseed seeds,mainly in the stem length,the fresh weight of the ground part,the leaf chlorophyll content was significantly decreased,and the accumulation of proline content in the leaves increased.4.A natural population containing 368 inbred lines of Brassica napus were used to identify the salt tolerance in seedling stage,mainly inspected the traits of root length,shoot length,shoot fresh weight and shoot sodium ion concentration and different traits of salt tolerance index was calculated.These traits have a wide range of phenotypic variation,exhibited the typical characteristics of quantitative character,and there was significant phenotypic correlation between some traits.All the traits of the heritability were more than 0.8,suggesting that these traits phenotypic variation were mainly controlled by genotype,and genetic characteristics were relatively stable.5.The genotype of the group was successfully identified by 60 K SNP chip in the previous study.Quality control of SNP markings according to the following criteria: the call rate was 0.7,SNP call frequency> 0.75;minor frequency> 0.05;AA,BB frequency> 0.03;and Gen Train score> 0.5.Finally,specific sites and positions with unclear loci were removed,and 19167 high quality SNPs were finally obtained for association analysis.The population structure and genetic relatedness of 368 materials were calculated using SNP markers.The population was divided into two subgroups P1 and P2.P1 subgroup consisted of 105 materials,and 263 of P2 subgroups materials.The average relative relationship among the whole population was 0.045,which indicated that the genetic relationship among the groups was weak.6.Combined with genotype data,the Q model and Q+K model were used to analyze the salt tolerance index and shoot sodium ion content in the three environments.A total of 75 SNPs were significantly associated with salt tolerance index phenotype and above ground sodium content(P <0.0001),of which 24 SNP markers were repeatedly detected in multiple environments.According to the LD between markers,75 SNP markers were integrated into 25 QTLs,which could explain phenotypic variation of 4.21%-9.23%,respectively.7.Using the genomic annotation information of Brassica napus,38 possible candidate genes related to salt stress were found within 500 kb upstream and downstream of significant SNP markers.Additionally,38 possible candidate genes were identified in genomic regions associated with salt tolerance indices.Among these genes,7 possible candidate genes related to salt stress were identified for ST-RL within 500 kb of peak SNPs,they are genes which were the orthologous to the Arabidopsis genes HOS10,TMT2,JR1,P5CS1,CYSA,ITN1,and ERD6.For ST-SL,22 genes were identified within 500 kb of peak SNPs;they are genes which were the orthologous to Arabidopsis genes ITN1,PYK10,PIP1 C,SAP18,MYB47,TIP2,HOS10,and PIP2 B.For ST-SFW,nine genes were identified within 500 kb of the peak SNPs,they are genes which were the orthologous to the Arabidopsis genes OSM34,RABG3 E,CDPK1,AOC2,DDF1,APA1,GDH2,PGIP2,and MAPKK2.These genes fell into several functional groups that are associated with plant salt tolerance,including transcription factors,aquaporins,transporters,and enzymes.The discovery of these genes will lay the foundation for the further cloning of the genes controlling the salt tolerance of rapeseed. |
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