Analysis Genome,Transcriptome Sequencing Of Stylo Colletotrichum Gloeosporioides And The CgALS Gene Of Pathogenic

Colletotrichum gloeosporioides?Penz.et Sacc.?could infect,such asrubber,mango,banana,sisal,Stylo,etc.many tropical economic crops,causing serious anthracnose,a serious threat to tropical agricultural production.In our country,Colletotrichum gloeosporioides were the main pathogenic bacteria of Stylosanthes anthracnose,disease occurs when production has been seriously affected,sometimes resulted in the death of a large area,and even crops.In this study,the use of a new generation of high-throughput sequencing platform?Ion Torrent?PGM?,Illumina HiSeq 2000/2500/MiSeq?for the high virulence strain CH008 genome,the bottom transcriptome was sequenced,assembled,splicing and function annotation,and whole genome sequence based on CH008,7 strains of different types,different pathogenicity isolates were re sequencing,and comments in detail comparative analysis.The main results are as follows:1,the Colletotrichum gloeosporioides pathogenic determinationThe research results show that the strain,pathogenicity of preservation of the decline,but on the whole,the high pathogenic strain always maintain a high pathogenicity.From the disease index can be determined,the whole genome,transcriptome sequencing the bottom with Stylosanthes anthracnose pathogen CH008 B type high pathogenic strains,re sequencing using CH010,CH013,CH020,CH450 for B type low pathogenic strains,CH247 type A highly pathogenic strains,CH036,CH100 for A type low pathogenic strains.2,Sequencing and analysis of complete genome of strains CH008The use of Ion Torrent?PGM?and Illumina HiSeq 2000 sequencing platforms were completed in CH008 strain small fragment library?400bp?and the large library?3K?of the whole genome sequencing,sequencing data were obtained using 3.1G and 4.3G respectively,the sequencing depth are respectively 50.35X and 68.89X,using Newbler and SOAPdonovo/Velvet completed the whole genome sequence assembly,assembly theresults showed the genome size is 57.21Mb,the Contig N50 156308bp,Scaffold N50 558940bp,compared with the whole genome sequence of Colletotrichum genus in the NCBI database,is higher than all the others,especially the Colletotrichum gloeosporioides?host grass mildew?genome 5.69 times and 4.95 times,indicating genome assembly results better,have more abundant biological information.Comprehensive utilization method of homologous notes and de novo annotation of gene prediction on the genome,predicted 37206 genes,through GO,COG,KEGG and other database enrichment and alignment,completed 24904 genes annotation,accounting for the prediction of gene 66.94%,also won the 4355 genes related to 507 KEGG pathways.In the PHI database,5950 genes associated with host interaction were also obtained.3,CH008 strains,the transcriptome sequencing and analysisUsing MiSeq Illumina sequencing platform to complete the sequencing of the CH008 strain of the transcriptome,obtained 6.1G sequencing data.Predicted 25379 unigenes,through COG,KEGG,GO and other database enrichment and comparison,completed the 16684 gene annotations,the total number of 65.74%of the total transcription of the gene.At the same time,the SSR analysis was carried out in 2609 genes,3242 potential SSR sequences,and 1505 pairs of SSR primers were designed,and 20 pairs of primers were selected for PCR amplification.The results showed that the SSR analysis results were good,and can be used to study the genetic variation of the pathogen in the column.4,completed in 7 strains of Stylosanthes anthracnose pathogen strains re sequencing and analysisBased on the whole genome sequence of CH008 strain,strain sequenced using Illumina HiSeq 2500 sequencing platform completed 7 strains of different types,different virulence,effective data were obtained by 12.09G analysis,quality control,sequencing data of good quality,re sequencing the genome of the sample average coverage depth is about 22X,the genome coverage is about 85%.Through the SNP,Small InDel,SV mutation analysis,were obtained for 2274884 of total SNP,transition and transversion ratio is about 2.33;the total InDel is 181999,the coding region was 20492;the total SV is 16042,also has carried on the detailed analysis and comment on the variation between the 7 samples.By clustering analysis,the high virulence strains were clustered into a group of people.It can be inferred that there are obvious differences between the high pathogenic bacteria and the low pathogenic bacteria.5,According to the completed sequencing database,1 screening models were set up to screen out the suspected disease related genes,and the suspected target gene CgALS?gene₁6548?was determined according to the information of the genome annotation.The use of knockout Split-marker gene,CgALS gene knockout,the results showed that CgALS gene knockout strains after pathogenicity or reduce the loss,slow growth rate,sporulation significantly reduced,preliminary studies suggest that CgALS is associated with pathogenicity.The study of Stylosanthes anthrax bacteria in the genome level analyses and basic research,and makes a preliminary study on genes associated with,which can deeply understand and excavate new key pathogenic genes,lay a good foundation for the study of Stylo anthracnose genetic variation and pathogenic mechanism etc..