Screening And Metabolic Characteristics Of Acetoin Production Strain TYF-LIM-RU47 Using Lignocellulose

Acetoin is an important four-carbon platform compound that occurs naturally in fruits or fermented foods,such as strawberries,cocoa,butter and white wine vinegar.Although it is mainly used as a food additive to enhance the flavor of products,Acetoin is also used to improve the aroma of other industrial products such as soaps,detergents and emulsions,and also because of its unique aroma,it is used in the food and cosmetics industries.The flavor enhancer and fragrance are also important metabolites secreted by microorganisms.Strain Rummeliibacillus sp.TYF-LIM-RU47 is a thermophilic and acid-resistant wild strain isolated and screened from Shanxi old vinegar by the research group.The 16 s r DNA of the strain was subjected to BLAST comparison at the interface boundary in the NCBI website,and identified as Rumelliella.The characteristics of the strain were further observed through basic biochemical experiments.It grows well in the base,and the main metabolite is acetoin.In order to deeply understand the metabolic mechanism and characteristics of the strain,the whole genome and transcriptome sequencing analysis of the strain was performed in this study.The acetoin-producing mechanism and metabolic pathway were determined from the genome annotation results.The transcriptome data was found by comparison Three up-regulated genes in lignin degradation pathway-related enzyme biosynthesis.Based on the advantages of the strain TYF-LIM-RU47 using lignocellulose,the optimal conditions for high-yield acetoin were obtained by optimizing the fermentation conditions.In addition,the strain can also use waste distiller's grains for growth metabolism,and simulate fermentation after adding fermentation broth Environment,the content of acetoin in the distilled product is significantly increased,and the taste is mellow.The main findings are as follows:1.After screening 5 different batches of vinegar samples,the 16 s r DNA of the 30 strains(bacteria)screened were subjected to homology comparison on the NCBI website,and the target strain TYF-LIM belonging to the genus Rumeliella-RU47.After basic physiological and biochemical experiments,it was concluded that the strain TYF-LIM-RU47 is a Gram-positive bacterium and has the characteristics of high temperature and acid resistance.2.According to the whole genome sequencing analysis of the strain,the genome size of the strain was3.30 Mb,the GC content was 37.44%,14 scaffold and 14 conting,and a total of 3418 genes were annotated.In the NR database results,we determined that the strain belongs to Lumei In the CAZy database,we found related enzymes that degrade cellulose and hemicellulose,and the presence of laccase suggested that the strain has the potential to degrade lignin.According to the existing metabolic pathway of acetoin,the important enzymes in the metabolic pathway of the strain were found through KEGG annotation information,and the existence of the gene was confirmed by next-generation sequencing,which confirmed the ability of the strain to produce acetoin.3.After screening of the fermentation substrate,it was found that the strain TYF-LIM-RU47 has a strong ability to utilize hemicellulose,and then xylose was used as a substrate.Through fermentation optimization,the concentration of xylose was 25 g / L,and the initial p H was 6.5.At 45 ?,200 r / min and 5%(V / V)inoculation volume,the best yield of acetoin can reach 15.8g / L.When lignin was used as a substrate,100 mmol / L of guaiacol was detected,and it was speculated that the strains could also use lignin.Using waste distillers grains as fermentation substrate,the yield of acetoin was as high as 30.8g / L.After simulating the brewing conditions,the optimal ratio of the substrate was found,and the content of acetoin in distilled liquor was up to 580 mg / L.4.In order to further investigate the transcriptomics characteristics of the strain TYF-LIM-RU47,a comparative analysis of the transcriptome sequencing results of the 17 h and 66 h fermentation samples using lignin as the main substrate,we found a total of 1049 differential genes Of which 695 genes are up-regulated and 354 genes are down-regulated,Through GO enrichment analysis of differential genes,we found that the enrichment of protein or peptide anabolic pathways and structural molecular activities is relatively significant.In the results of enrichment analysis of differential gene KEGG,the biological effects of enzymes related to lignin degradation Analysis of the synthetic pathways revealed a total of 3up-regulated genes.This part of the study can provide a theoretical basis for exploring the lignin degradation pathways.