Genome Assembly And Phylogenetic Analysis Of Cell Organelles In African Cultivated Rice

As an important part of plant genetic information,chloroplast genome and mitochondrial genome play important roles in the research of phylogenetic evolution,species identification,nuclear-cytoplasm interaction,and genetic engineering.The African rice(Oryza glaberrima Steud),one of the two common rice,is an important rice germplasm resource.The assembly of the organelle genome of African rice could be completed through whole genome sequencing,which is of great significance for broadening the genetic diversity of rice germplasm resources and innovating rice breeding approaches.Up to present,rice varieties that have been sequenced include common cultivated rice and some wild rice varieties,however,no report has been made on the mitochondrial genome sequence of African rice RAM3.In this study,based on the sequencing data of multiple genomic libraries constructed from African rice RAM3,the assembly and analysis of plant organelle genomes were attempted.The main results are as follows:1.Assembly and analysis of chloroplast genome: using the Miseq PE2500 double-end sequencing data,using the 27 chloroplast genome sequences collected by NCBI as reference,the African rice chloroplast genome sequence was extracted through BWA comparison,and initially assembled with Discovar Denovo software and obtained The LSC,SSC and IR sequence fragments were further linked to complete African rice RAM3 chloroplast genome.The chloroplast genome of African rice RAM3 is 134582 bp in length,showing a typical four-segment structure,encoding 117 genes and containing 83 protein-coding genes,4 r RNAs,and 30 t RNA genes;its genome structure,gene sequence,content,and codon usage Similar to other published rice chloroplast genomes.A total of 32 scattered repeats and 32 tandem repeats were detected in the chloroplast genome of African rice RAM3.Sixteen simple sequence repeats(SSR)analysis of African rice RAM3 chloroplast genome revealed that SSRs are poly-A or poly-T and are not uniformly distributed within the chloroplast genome.2.Comparative genomics study:While there was an overall similarity between the chloroplast genomes of African rice RAM3 and the five other rice genera AA genome species(Japocin?Indica?Nivara?Rufipogon?Barthii),greater variation was found in the intergenic region;The IR border genes of African rice RAM3 were similar to that of the 5 AA rice chloroplast genomes,though slightly different in their distance?; based on 7 different chloroplast gene datasets of 31 plant species,a phylogenetic tree was constructed and phylogenetic analysis was performed.It was found that the LSC,CDS,full sequence,and IGS datasets of the chloroplast genome were in evolution.The support rate in the analysis is high and the result is more reliable.Five chloroplast-specific markers of Africa rice were screened to confirm that all the introgression lines developed in this laboratory had African rice cytoplasm.3.Mitochondrial genome assembly and analysis:The mitochondrial genome of African rice RAM3 was spliced and assembled using three softwares The assembly sequence was 495258 bp in length and encoded a total of 84 genes containing 64 protein-coding genes,3 r RNAs,and 28 t RNA genes.The entire mitochondrion contains 16 introns distributed in nine protein-coding genes.A total of 94 scattered repeats and 29 tandem repeats were detected in the mitochondrial genome of African rice RAM3.Thirty-seven simple sequence repeats(SSR)analysis of the mitochondrial genome of African rice RAM3 revealed that: SSRs are poly A or poly T,similar to the chloroplast genome,and they are not uniformly distributed within the mitochondrial genome.Analysis of codon usage bias revealed that leucine has the highest probability of use,and tryptophan and methionine are the least.4.Analysis of the intergenic sequence of African rice RAM3 mitochondrial genome revealed a total of 75 AT enrichment regions.The maximum difference of A+T can reach 35.19%;the secondary structure of 28 t RNAs in the mitochondrial genome of African rice RAM3 is predicted,and 58 mismatches and 1 deletion are found;Analyze the gene selection pressure and find that gene nad6 has the highest selection pressure and gene nad2 is under the lightest selection pressure.Gene cob is selected under neutrality.5.Analysis of the sequence shifts between African rice RAM3 organelle genomes revealed that the sequences moved from chloroplasts to mitochondria and the total fragments were 22311 bp in length,accounting for 4.5% of the total length of the mitochondrial genome of African rice RAM3.A total of 21 migrated fragments were greater than or equal to 100 bp in length,with the longest fragment being 6752 bp in length.