Molecular Mechanism Analysis Of A Root Tip Lethal Mutation In Brassica Napus By RNA-seq

Brassica napus(L.)is one of the important oil crops among the three major types of oil seed rape.Brassica napus(2n=38,genome AACC)originated from natural interspecific hybridization between B.oleracea(2n = 18,genome CC)and B.rapa(2n=20,genome AA)and then form heterologous polyploids at about 7500 years ago.Study in cytology and molecular genetics of rapeseed breeding have made significant progress now.However,the molecular genetics and breeding but the molecular mechanism of root growth and development not fully elucidated yet.Transcriptome based rape root development will be helpful to improve the quality of rapeseed breeding.In this study,we developed the transcriptomic profile of root apical,different stages of root tip development followed by the gene expression analysis of different stages of apical development.The main findings are as follows:1.Microscopic observation of root tip development at different stages immediate after the seed germination by electronic microscope revealed remarkable difference between mutant and normal roots.Furthermore,a black spot,was appeared first on the mutant root tip that increases gradually,and ultimately stopped the development of root.2.We used Illumina Hi Seq 2000 two-terminal second-generation sequencing technology for transcriptomic profiling of normal and mutant root apical.And found 44.5GB of transcriptome data.We used Hisat2 software for making comparison.Hisat reads ratio about 90%.3.We analyzed the database of two developmental stages of apical normal and mutant.Results indicated that a total of 47,968 genes are expressed in eight samples.the expression of different genes in the first stage was significantly higher than the second stage.Moreover,from a Total expressed of 3,311 genes 1,864 genes were up-regulated while 1,447 were down-regulated.On the other hand it was noted that 425 genes are up-regulated and 488 genes are down-regulated.in the second stage.We investigated the GO classification,GO enrichment and KEGG pathway of the genes.In the GO classification,it was deduced that.the genes are mainly distributed in the catalytic activity,cell and organelle generation,metabolic process,cellular process,regulation of biological process,response to stimulus and biological regulation.In the GO enrichment,the two genes were significantly enriched into two pathway that is nucleic acid binding pathway(GO: 0001071)and structural component of cell wall component binding pathway respectively.Two genes were compared to the KEGG,and two significant metabolic pathways were obtained that is Steroid biosynthesis pathway(KO: ko00100)and,alpha-Linoleic acid metabolic pathway(KO: 00592).4.Go annotation analysis revealed that four different genes that are involved in cell killing are Bna A05g03400 D,Bna C04g49050 D,Bna A02g17520 D and Bna C02g23400 D.We compared these four genes to the Arabidopsis database and found that Bnaco2g23400 and Bna A02g17520 were homologous genes.Whereas,Gene(Bna A02g17520D)show homology with Arabidopsis thaliana(ATG37870)that is responsible for phosphoenol pyruvate carboxylase.It is key enzyme for gluconeogenesis and is affected by MC9-dependent protein activity as a substrate for MC9 and has a function of cell death-related degradation.The GO function of these four genes is related with our phenotypic traitsWe enriched a pathway(Protein processing in endoplasmic reticulum Ko0441)related to endoplasmic reticulum degradation of the metabolic pathway from the two mutual validation of GO enrichment score.