Transcriptomics Study And SRNA Difference Analysis In Each Growth Phase Of Streptococcus Thermophilus MN-ZLW-002

With the enhancement of consumers' health concept,fermented dairy pro ducts represented by yogurt had become more and more popular.By the end of 2018,the sales volume of China's yogurt market had exceeded 140 billion RMB.As one of the most important fermentation strains of dairy products,the market demand for Streptococcus thermophilus had also increased.In the preparation of industrial starter,the design of each nutrient component of high-density medium was the key to obtain optimal biomass.Streptococcus thermophilus was an auxotrophic strain who had extremely demanding for various nutrients in the medium,especially the three main components of carbon source,nitrogen source and nucleic acid.In the fermentation process,the study of the metabolic mechanisms of these three components was the key to optimizing the components and fermentation parameters of Streptococcus thermophilus high-density medium,however,the research on this part was currently lacking.Genes expressed at transcriptional level and s RNA expressed at noncoding RNA level in Streptococcus thermophilus MN-ZLW-002,as the experimental strain forthis study,at different growth stages in chemical synthesis medium were determined by high throughput sequencing technique.In this study,1937 genes were obtained by transcriptomics sequencing,and functional(GO)annotation and metabolic pathway(KEGG)annotations were performed.Based on the difference in the expression levels of genes in each sample,genes differentially expressed at each growth period were screened for functional enrichment and metabolic path enrichment analysis.This study focused on the global transcription of nucleic acid metabolism,carbon metabolism,and amino acid metabolism during fermentation.In nucleic acid metabolism,the de novo synthesis pathway mainly occured in the log phase,in which the pur operon was up-regulated more than 5-fold in the logarithmic pre-stage and about 2-fold in the late logarithmic expression.The pyrimidine de novo synthesis pathway was in a low expression state throughout the growth period,however during the stationary phase,the expression of the pyr operon was slightly upregulated.In the carbon metabolism,galactose metabolism mainly occurs in the logarithmic phase,and the two ke y genes lac S and lac Z are up-regulated by 1.2-fold in the logarithmic pre-stage and up-regulated in the late logarithmic phase by 3-fold.However,during the lag phase,galactose metabolism is inhibited,an d the expression of both genes was down-regulated by more than 1 fold;Furthermore,gal K encoding galactose kinase was highly expressed throughout the growth period,particularly in the lag phase and log phase,indicating that Streptococcus thermophilus MN-ZLW-002 might have the ability to metabolize galactose.Glycolysis was an important energy supply pathway in the fermentation process.The lag phase is the most active period of glycolysis in which the key genes glk,pyk and pgk were up-regulated by more than 5 times.However in the log phase and the stationary phase,gene expression in this pathway was down-regulated to varying degrees.As a product of glycolysis,pyruvate showed that the reaction of pyruvate to acetic acid mainly occurred in the lag phase and the logarithmic pre-stage,and the expression of ldh catalyzing the reaction was up-regulated by 1.6-fold and 0.2-fold,respectively.The reaction of pyruvate to acetyl-Co A occurred mainly at the end of the logarithm,and the pbh BCD catalyzing the reaction was up-regulated by 1.9-fold in the late logarithmic phase.Fatty acid metabolism mainly occurred in the lag phase and the logarithmic phase.During this process,the rate-limiting gene acc B was up-regulated by 2.9-fold in the lag phase,but did not change in the logarithmic phase.However,the exp ression in the logarithmic pre-stage and the stationary phase showed a downward trend.In the amino acid metabolic pathway,differentially expressed genes were mainly enriched in glutamate,alanine,aspartic acid,arginine,glycine,serine,cysteine,methionine,proline,leucine and isoleucine metabolic pathway,and the metabolism of these amino acids also showed differences in different growth stages.The lag phase was the most active period of amino acid metabolism,and almost all genes with different cha nges were up-regulated;In the log phase,the main reaction was the conversion of aspartic acid to fumaric acid and oxaloacetate,the conversion of glutamic acid to succinic acid,glucose 6-phosphate,ribosyl 5-phosphate and carbamoyl phosphate,and the conversion of serine to Pyruvate and tryptophan,methionine was converted to N-formyl-L-methionine,arginine acid deiminase pathway(ADI);the main reaction occurring during the stationary phase is glutamate production Glucose 6-phosphate,serine was converted to pyruvic acid,and dihydrolipoamide-E was converted to lipoic acid amide-E in the branched-chain amino acid metabolic pathway.A total of 530 candidate sRNAs were identified,including 198 asRNAs,135 sRNAs from intergenic regions,and 197 sRNAs from untranslated region(UTRs).Target genes of s RNA were predicted by Inta RNA,and 390 target genes were enriched into 99 metabolic pathways.Analysis of the differential expression of s RNA showed that 238,83,194 and 139 sRNAs significantly changed during each growth period.Further analysis of the 14 most significant sRNAs from each growth period revealed that the major metabolic pathways for enrichment of these potential target genes involved carbon metabolism,amino acid biosynthesis,ABC transporters,ami no sugars and nucleotide sugars.Metabolism,purine metabolism and phosphotransferase systems.A preliminary analysis of the potential regulatory mechanisms of these sRNAs was performed in conjunction with the expression of these target genes in transcriptomic data.The sRNAs differentially expressed during the lag phase had a positive regulatory effect on the potential target gene s;In the logarithm pre-stage,some sRNAs might play the role of the housekeeping genes,and control the intracellular metabolic flux by negatively regulating the target genes;At the late logarithm,the differentially expressed s RNA negatively regulated the target genes,and the detachment inhibited the metabolic activity of the strain,ensuring the stable survival of the strain i n a complex growth environment;During the stationary phase,6S RNA acted to maintain cell homeostasis by positive regulation of pyrimidine metabolism.