Genetic Mechanism Controlling Wax Trait On Leaf Surface Of Brassica Napus Seedlings

Rapeseed(Brassica napus L.)is an important source of edible vegetable oil,animal feed and biofuel.Biotic and abiotic stresses such as drought,low temperature,pests and diseases seriously endanger the growth and development of rapeseed seedlings.Cuticular wax is the first barrier of above-ground tissues of many plant species against adversities.The chemical composition and crystal structure of cuticular wax determine the physicochemical properties of the plant surface.Revealing the molecular mechanism controlling leaf wax formation is of great significance to the genetic improvement of stress resistance(tolerance)in rapeseed.In this study,a collection of 297 core accessions of B.napus was used as research material.Genomewide association study(GWAS)revealed single nucleotide polymorphisms(SNPs)significantly associated with leaf wax coverage of rapeseed seedlings.Genomic analyses combined with transcriptome uncovered candidate genes controlling wax trait in rapeseed.The main results are as follows:1.The degree of wax coverage could be judged by the significant differences in leaf color of rapeseed seedlings,so that the wax phenotypes of B.napus accessions could be classified into low-wax,medium-wax and high-wax types.Scanning electron microscopy showed that the density of epicuticular wax crystals was determined by the leaf wax content.High light treatment significantly increased the amount of wax deposited on the leaf surface of rapeseed seedlings.The resistance against Sclerotinia sclerotiorum infection of rapeseed accessions with high-wax phenotype was significantly higher than that of low-wax accessions,and the resistance of rapeseed leaves to S.sclerotiorum was significantly reduced after removing the epidermal wax.The average fresh weight of above-ground parts of rapeseed seedlings with high-wax phenotype was significantly lower than that of low-wax accessions.2.Based on the phenotypic variation of wax trait at the seedling stage of 297 rapeseed accessions,89 SNPs significantly associated with leaf wax coverage were identified by GWAS.Transcriptome analysis of rapeseed accessions with contrasting differences in leaf wax content integrated with GWAS results revealed 17 differentially expressed candidate genes such as Bna A02.LOX4.Selective sweep analysis uncovered the genome-wide selective traces between high-and low-wax populations.A total of349 candidate genes with significant selective signatures were identified by the crossanalysis of selective traces and differentially expressed genes between high-and lowwax accessions,which were further used to briefly speculate the wax biosynthetic pathway in the leaf epidermis of rapeseed.3.We found a significant correlation between the geographical distribution patterns of different wax phenotypes and the rapeseed ecotypes.Most winter accessions originating from Germany showed the high-wax phenotype,while semi-winter accessions distributed in China were mainly the low-wax phenotype.The allelic variants in the candidate genes,Bna Cnn.CER1,Bna A02.CER3,Bna C02.CER3 and Bna A01.CER4,were the potential genetic basis for the divergence of wax phenotypes among different ecotypes of rapeseed.This study revealed the association between wax trait and ecotype in rapeseed.The genetic mechanisms underlying the divergence of wax phenotypes on the leaf surface of rapeseed were analyzed at the genomic and transcriptomic levels,which identified a number of genetic loci and candidate genes significantly associated with leaf wax content.These results provide germplasm resources and theoretical bases for rapeseed breeding.