Transcriptomic Analysis Of Rice In Response To The Infection Of Different Blast Fungus

With the development of biological science and technology,the emergence of genome high-throughput sequencing technology,also known as "next generation"sequencing technology and third-generation sequencing technology of the genome,makes it possible to perform a detailed analysis of the transcriptome and genome of a species.Transcriptome sequencing has become the main method for studying gene expression.The regulation of transcription level is currently the most studied and the most important regulation method of organisms.In this paper,3 Near-Isogenic Lines(NILs),including CN1,CN-4a,and CN-4b,which were developed from parent line C039 and showed resistant ability to rice blast,were used as materials.Whilst,two blast fungus race,GUY 11and 81278ZB15 with different pathogenic ability,were used to infect rice seedling.The second-generation sequencing technology was applied to sequence transcriptomes(RNA-seq)of the 3 test rice varieties at 24h and 48h after blast fungus inoculation.The results were shown below.1.CN1,CN-4a,and CN-4b all resist to the infection of rice blast fungus 81278ZB15,while only CN-4b resists to GUY11.Analysis of the RNA-seq data showed that the high-quality sequencing data of all the samples was 92.74-95.41%of its raw data.TopHat tool was applied to align the high-quality sequencing reads to the reference genome,which was downloaded from http://www.msu.edu/genome.Totally,there were 76.8%and 85.2%sequencing reads mapped to the reference genome,among which more than 74.5%were exons.Moreover,all the RNA sequences were evenly distributed in the 5'-3' direction of the genes.The results indicated that the transcriptomic sequencing data with high quality could be used for further analysis in the next research.2.The significantly differential expressed genes(DEGs)in the 3 test rice varieties under the infection of 2 different blast fungus were obtained using Cuffdiff tool withthe threshold value of p_value<0.005 and q_value<0.05.The result demonstrated that there were more DEGs in all the test rice variety at 24h after inoculation than at 48h after inoculation.CN-4b had 7026 DEGs at 24h after inoculation,which was significantly higher than the other 2 test varieties,2808 and 2586.The GO annotation pathway of rice CN-4b up-regulated DEGs at 24h was significantly different from the other 2 test varieties,and was mainly enriched in translation and gene expression.3.The above RNA-Seq results indicated that LOC_Os08g13465 in CN-4b significantly induced by GUY11 than the other 2 test varieties.This result was fuether validated by using qRT-PCR.Whilst,CRISPR/Cas9 experimental results showed that LOC_Os08g13465 had the resistant ablity to Guy 11 infection.We searched the transcription factors for LOC_Os08g13465 against transcription factor database,and found two factors,TRABland OSBZ8.Moreover,the expression abundance of TRAB1 in CN-4b after GUY11 infection 24h was significantly higher than the other2 test rice varieties.These results showed that LOC_Os08g13465 was activated in CN-4b by transcription factor TRAB1,which was induced by Guy 11 infection.In summarry,the results in this paper provide new insight into the resistant ability of rice under GUY11 infection and implicate a new blast-resistant gene for development program of new rice varieties.