Transcriptome Study Of Paenibacillus Mucilaginosus KNP414Based On RNA-Seq Technology

Paenibacillus mucilaginosus, a critical silicate bacterium in the biogeochemical cycling ofpotassium, phosphorus, and other soil elements, is able to degrade insoluble soil minerals withthe release of nutritional ions and fixes nitrogen. These functions make P. mucilaginosus widelyused in agriculture as a biofertilizer, in bioleaching as a biocomplexing agent, and in wastewatertreatment as a bioflocculant. However, these mechanisms keep unknown. RNA-Seq technologyprovides an efficient and accurate way to study transcriptome. In order to know about thenitrogen metabolism, we present the qualitative and quantitative RNA-Seq analysis of thetranscriptomes of P. mucilaginosus KNP414at log-phase and stationary-phase cultured innitrogen and nitrogen-free medium, respectively.Sample the cells at11h and18h as the exponential phase and stationary-phase undernitrogen and nitrogen-free medium, respectively. The total RNA was extracted, the cDNA librarywas constructed, and the transcriptomes was sequenced using Hiseq2000. We analyzed the dataof two18h samples (18hN and18hN+). The original sequencing data of the two samples was1.77G and3.48G, respectively. After removing the adapters and reads with the mean quality lessthan20,17,298,505and34,275,841high-quality reads were obtained. These high-quality readswere normalized and utilized to calculate FPKM (Fragments Per Kilobase of transcript perMillion fragments mapped) through matching the genome. Altogether7457and7765out of7811predicted genes in the genome of KNP414were expressed in the two samples, respectively.As the results, there were895upregulated genes, and4596downregulated genes in the contrastsof18hN versus18hN+.The GO enrichment analysis of differentially expressed genes was carried out on the base ofthe GO database, and the statistical significance was evaluated by Fisher’s exact test in thepackage Blast2GO. There were42statistically significant GO teams in the upregulated genes,and80statistically significant GO teams in the downregulated genes in the group of18hN versus18hN+. Pathway enrichment analysis was subsequently carried out using the KOBAS2.0based on KEGG database with hypergeometric test. The statistically significantly enrichedpathways were identified in differentially expressed genes compared with the whole genome.According to the results, there were3statistically significant upregulated pathways, and3 statistically significant downregulated pathways. This analysis revealed significantly over-andunder-represented terms, which highlighted the process and functions contributing to specificphenotypic traits of the P. mucilaginosus KNP414under different growth conditions.The related metabolic pathway (nitrogen metabolism, amino acid metabolic process, proteinmetabolic process, spore germination, transport, and signal transduction and regulation) of thedifferentially expressed genes was especially analyzed in18hN and18hN+.10out of32predicted genes related to nitrogen metabolism, which might be involved in nitrogen-fixtion,were upregulated in18hN. Besides, genes related to capsular polysaccharide biosynthesis andspore germination were overexpression in18hN. The expression of genes in18hN+wasabundant. There was an upregulation of genes resistant to the change of physicochemicalproperty as well as nitrogen deficiency. Totally75.62%(152/201) of pridicted amino acidmetabolic process related genes were upexpressed in18hN+compared with11hN+. At the sametime, many genes associated with protein synthesis through all the complex process wereupexpressed, including RNA polymerase, aminoacyl-tRNA biosynthesis, ribosome, peptidesynthesis, protein transporter, protein catabolic process, and protein folding and modificationprocess. In addition, genes related to ABC transporters, two-component and phosphorelay signaltransduction systems were all induced.In summary, the transcriptomes of P. mucilaginosus KNP414under nitrogen andnitrogen-free were analyzed using RNA-sequencing method, which is the basis to know moreabout the nitrogen metabolism. However, no differentially expressed genes were found directlyrelated with nitrogen fixation. Therefore, further research is necessary to clarify the mechanismof nitrogen fixation for P. mucilaginosus KNP414.