Screening And Genomic Analysis Of A Lignocellulose Degrading Bacterium

A lignocelluloses-degradation bacterium,referred to S12,was isolated from the soil.The biological characteristics and genetic characteristics of this isolate were identified by analyzing its whole genome and related gene expression.The corresponding culture was collected under different temperature and carbon sources.This study provides the genetic background and biology mechanism of lignocellulose degradation processes within bacteria.Soil sample was collected from Mt.Zijin,Nanjing.Three kinds of lignin analogues(Azure-B;Phenol red;Guaiacol)bleaching/dyeing method were applied to separate bacterium with ability to degrade lignocellulose.The classification of the species was based on the combined analysis of 16s rRNA gene and genome sequences.Ultraviolet spectrophotometric method was used to determine the enzyme activity of manganese peroxidase(MnP),laccase(Lac),carboxymethyl cellulose enzyme(CMCase)and filter(FPA).The genome of strain S12 was sequenced on Illumina Miseq and 454 GS Junior sequencing platform.The whole genome was assembled by Newbler,and subsequently,blasted against COG and KEGG database.Some important enzymes were further validated by quantitative PCRs.Finally,the whole genome was submitted to Genbank,with CP010557 as its asseccion number.As the results,the genome of individual bacterium S12 was composed of 5,522,044 basepairs,and its G+C content was 57.47%.On both 16S rRNA and genome trees,the species was most closely related to Raoultella ornithinolytica.In the lab experiments,it took 28 h to grow to the plateau stage in the liquid CMC-Na medium.The activities of the enzymes related to cellulose degradation were peaked at that time too.Bioinformatics analysis showed that the strain S12 had genes encoding the key enzymes in the lignin degradation pathways,such as peroxidase,hydrogen peroxide-producing enzymes,catechol 1,2-dioxygenase and protocatechuate 3,4-dioxygenase,cellulose synthase and ornithine carbamyl acyltransferase.Comparing with glucose,the expressions of those genes are significantly higher when it cultured in lignin monomer compounds,such as syringic acid or gallic acid.According to the analysis of the KEGG pathway,strain S12 has complete pathways of cellulose degradation and ethanol generation.In addition,strain S12 has the protein-coding gene cluster within bacterial type ? secretion system,which is proved to be the most important system of extracellular degradation in gram-negative bacteria.Meanwhile,one unclassified gene on strain S12 was predicted to be carbon storage regulation factor A(Carbon storage regulator,CsrA),involved in the degradation of lignocellulose and the following small molecules.Our study provides the better insights on the mechanisms of soil bacteria to degradate the lignocelluloses.It has great significance in the development of lignocellulose application industry.