Preliminary Research On Formation Mechanism Of Stiff Stem In Brassica Napus

Lodging is one of the main reasons for reduction of rapeseed output and limitation of mechanized harvesting.Breeding of lodging-resistance varieties is one important goal for rapeseed.Dissection of lodging-resistance characteristics and mechanisms of stiff-stem rapeseed is very important to stabilize and improve the rapeseed output and the application of rapeseed mechanized harvesting.Brassica napus YG689 in this study is stiff-stem one from the hybrid of Capsella bursa-pastor is and B.napus ZY821.Analysis of morphology and anatomical structure,lignocellulose component and content,comparison of the transcriptomes,comparison of the metabonomics and QTL mapping for lignocellulose were performed on the stems of different development stages of YG689 and ZY821 as a control to reveal the stiff-stem related morphologic,genetic and metabolic bases of YG689 and preliminary uncover the machanism of stiff-stem formation of YG689.The major research results in this study were as following:?1?The stem stiffness of YG689 was significantly higher than ZY821,and lodging resistance of YG689 was higer than ZY821.The stem anatomical structure results showed that lower quantity and larger distribution of vascular bundles,more interfascicular fiber,and larger lignin distribution in YG689 were than that in ZY821,suggesting that the stem lignin content of YG689 was higher than that of ZY821.Comparison of stem lignocellulose component and content of YG689 and ZY821 in the seedling stage,budding stage,initial flowering stage,final flowering stage and mature stage showed that the lignin content,cellulose content,hemicellulose content and lignin monomer content of YG689 were higher than that of ZY821.These results indicated that stem lignocellulose charactersitics were related to the stiff-stem and lodging-resistance of YG689.?2?The stem transcriptome of YG689 and ZY821 in the initial flowering stage was investigated using RNA-seq technology,respectively.A total of 8,644 differently expressed genes?DEGs?were detected.GO enrichment analysis and Pathway enrichment analysis showed that DEGs were significantly related to regulation of plant-type hypersensitive response,and involved in Ribosome,Arginine and proline metabolism,Cysteine and methionine metabolism,etc.A total of 624 DEGs were not aligned to B.napus reference genome,and 108 of them could be aligned to C.rubella reference genome?6 DEGs with homologous rate ?80%?.GO and KEGG annotation of DEGs?with homologous rate ?40%?aligned to C.rubella reference genome showed that they were involeved in response to stress,suggesting that defense-related genes of C.bursa-pastoris were transferred into the rapeseed genome by intertribal crosses.These results demonstrated that the genes transfer of C.bursa-pastor is hybridated with ZY821 had generated new genes and thus formed the stiff-stem characteristics of YG689.30 DEGs between YG689 and ZY821 were closely correlation to lignocellulose biosyntheis.The expression charateritics of 12 DEGs in the stems of YG689,ZY821 and TN070?another control?of the budding stage,initial flowering stage and final flowering stage showed that the number of highly expressed DEGs in YG689 of the initial flowering stage was maxmum,and the high expression levels of BnaA09g42650 in YG689 were maintained to the final flowering stage.?3?The stem metabolomics of YG689 and ZY821 in the budding stage,initial flowering stage and final flowering stage was investigated using GC-TOF/MS and UPLC-QTOF-MS technologies,and 40,54 and 31 differently metabolites were identified,respectively.These differently metabolites were dominantly involved in glycolysis,TCA cycle,pentose phosphate pathway,galactose metabolism,phenylalanine,tyrosine and tryptophan biosynthesis,etc.The content of raffinose,cis-Aconitic acid and mesaconic acid showed difference between YG689 and ZY821 for all of the three development stages,suggesting that their accumulation might be related to the stiff-stem characteristic of YG689.?4?QTL mapping of lignocellulose content was performed in the KN DH population,and 60 identified QTLs were obtained.Identified QTLs for the same trait with overlapped confidence intervals were integrated into 52 consensus QTL using a meta-analysis method with BioMercator 4.2.Based on the collinearity relationship between the genetic linkage map and the B.napus reference genome,and differently alleles of the parents re-sequencing,5,522 candidate genes were identified in 52 QTL regions of lignocellulose.Further,a total of 130 homologous genes of A.thaliana,that were involved in the biosynthesis and regulation of lignocellulose,were identified within confidence intervals of 52 QTLs by Mapman software,and these homologous genes were used to construct a potential regulatory pathway controlling stem fibre biosynthesis.By integration of the DEGs,the different metabolites and fibre-related candiated genes in the QTL region,a linked metabolic pathway was predicted and included 14 different metabolites and 8 key enzymes,and these different metabolites and enzymes might affect stem lignocellulose biosynthesis of YG689.Collectively,the stem morphological and anatomical charateristics of stiff-stem YG689 included high stem stiffness,large distribution of lignin in the vascular bundles and interfascicular fibers,and high content of lignin,cellulose and hemicellulose.Multi-omics analysis showed that NewGene₂16?TPS7?,NewGene₆060?MAT3?,NewGene₃483?BGLU40?,BnaA02g18920D?TPS1?,BnaA09g42650?PRX17?,BnaC05g48040D?MS2?,and BnaC05g48570D?GAUT13?might be the candidate genes associated with the stiff-stem charateristics of YG689.The accumulation of raffinose,cis-Aconitic acid and mesaconic acid could be one of the factors for the stiff-stem formation of YG689.