Transcriptome analysis of gene expression of strains JNB5-1 and ZK66
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ABSTRACT: RNAseq analysis was performed to evaluate gene expression differences between strains JNB5-1 and ZK66. S. marcescens JNB5-1 and ZK66 cells were grown in LB medium for 12 h before harvesting. The collected cells were then treated with RNAprep pure Kit (TIANGEN) to extract total bacterial RNA and delivered to GENEWIZ in dry ice for transcriptome resequencing analysis. For annotation, the genome of S. marcescens WW4 (NC_020211.1) was used as reference. A total of 24048918 reads matched to the referenced genome in the sample of JNB5-1, and 24569696 reads in the sample of ZK66. The differentially expressed genes (DEGs) were determined between strains JNB5-1 and ZK66 with the standards of false discovery rate (FDR) ≤ 0.05, fold change |log2Ratio|≥1. Transcriptome data showed that expression of 641 genes were upregulated while 784 genes were down regulated by at least 2-fold when comparing the metR mutant strain ZK66 to its parent strain JNB5-1. Based on the annotation of KEGG_B_class, the up-regulated and down-regulated genes were classified into 30 and 27 major cellular processes, respectively. The transcriptome data indicated that MetR may regulate a variety of cellular processes in S. marcescens, including prodigiosin synthesis, and cell motility.
Project description:RNAseq analysis was performed to evaluate gene expression differences between strains JNB5-1 and SK68. S. marcescens JNB5-1 and SK68 cells were grown in LB medium for 4 h before harvesting. The collected cells were then treated with RNAprep pure Kit (TIANGEN) to extract total bacterial RNA and delivered to GENEWIZ in dry ice for transcriptome resequencing analysis. For annotation, the genome of S. marcescens WW4 (NC_020211.1) was used as reference. A total of 32420858 reads matched to the referenced genome in the sample of JNB5-1, and 32993470 reads in the sample of SK68. The significantly differentially expressed genes (DEGs) were determined between strains JNB5-1 and SK68 with the standards of p-value≤ 0.05, fold change |log2Ratio|≥1. Transcriptome data showed that expression of 210 genes were significantly up-regulated while 106 genes were significantly down-regulated by at least 2-fold when comparing the rcsB mutant strain SK68 to its parent strain JNB5-1. Based on the annotation of KEGG_B_class, the up-regulated and down-regulated genes were classified into 18 and 17 major cellular processes, respectively. The transcriptome data indicated that RcsB may regulates a variety of cellular processes in S. marcescens, including prodigiosin synthesis, and cell motility.
Project description:RNAseq analysis was performed to evaluate gene expression differences between strains JNB5-1 and SK8-37. S. marcescens JNB5-1 and SK8-37 cells were grown in LB medium for 10 h before harvesting. The collected cells were then treated with RNAprep pure Kit (TIANGEN) to extract total bacterial RNA and delivered to GENEWIZ in dry ice for transcriptome resequencing analysis. For annotation, the genome of S. marcescens WW4 (NC_020211.1) was used as reference. A total of 31206068 reads matched to the referenced genome in the sample of JNB5-1, and 29862970 reads in the sample of SK8-37. The significantly differentially expressed genes (DEGs) were determined between strains JNB5-1 and SK8-37 with the standards of p-value≤ 0.05, fold change |log2Ratio|≥1. Transcriptome data showed that expression of 60 genes were significantly up-regulated while 558 genes were significantly down-regulated by at least 2-fold when comparing the psrA mutant strain SK8-37 to its parent strain JNB5-1. Based on the annotation of KEGG_B_class, the up-regulated and down-regulated genes were classified into 10 and 19 major cellular processes, respectively. The transcriptome data indicated that PsrA may controls a variety of cellular processes in S. marcescens, including prodigiosin synthesis, serrawttin W1 biosynthesis, extracellular polysaccharide production, biofilm formation, cell motility and T6SS-mediated antibacterial activity.
Project description:We report the application of transcriptome sequencing technology for high-throughput profiling of Serratia marcescens for producing prodigiosin. By obtaining over 163 million bases of sequence from Serratia marcescens genome DNA, we generated transcriptome -state maps of Serratia marcescens 12h cells, 24h cells, and 36h cells at 30C and 37C,respectively. We explored the mechanism of S. marcescens response temperature regulation at the transcription level through transcriptome sequencing technology. We found that the pig gene cluster at low temperature would favor at the transcriptional level, however, higher temperature resulting in instability and loss of enzyme activity. Numerous amino acid metabolic pathways involved in prodigiosin biosynthesis in S. marcescens responded to temperature changes, and metabolic fluxes were directed towards prodigiosin biosynthesis. At the same time, quorum sensing, two-component regulatory system and sRNA were stimulated by temperature to regulate PG biosynthesis and involve strain virulence and exclusive genes. Moreover, inhibition factors was the one reason for S. marcescens incapable synthesis of prodigiosin at 37C. This study laid a good foundation for understanding the biological functions of prodigiosin, improving the temperature tolerance of industrial strains, and excavating temperature-sensitive regulatory elements.
Project description:Seeds were germinated on 1/2 MS plates plus 1% sucrose and placed at 4°C for 3 days and then transferred to white light for 12 h before placed in blue light for another 4 days. Total RNA was extracted from samples including WT, agb1, cry1 cry2 and hy5 with RNAprep plant kit (TIANGEN), and followed by DNase I (Takara) treatment. This study reveals the molecular basis for light and G-protein crosstalk by analyzing gene expression profile under control of both signals.
Project description:The acid tolerance of industrial strains is a significant challenge in the fermentation process. The bacterium Serratia marcescens is part of the Enterobacteriaceae family of eubacteria, which is a potential industrial microorganism. However, the molecular mechanism behind S. marcescens acid resistance is not properly understood. In this study, we screened for novel regulators that respond to acidic conditions by a Tn5G transposon insertion mutagenesis of S. marcescens and found mutations in a gene encoding for the HTH_XRE super-family regulatory protein member, here named xrpA. Using transcriptomics and further experiments, we showed that the xrpA disruption conferred pleiotropic phenotype changes, including highly decreased biomass, altered cell shape, H+-ATPase activity, and deficiency of cell membrane permeability and integrity, compared with those of the parent (JNB5-1) strain at low pH. These data revealed that the molecular mechanism by which xrpA affects acid resistance of S. marcescens is through positive regulation of cell membrane permeability, integrity, and H+-ATPase activity to maintain intracellular homeostasis at low pH. Finally, we constructed an acidic resistant strain JNB5-1/pSX1314 by overexpression of xrpA in S. marcescens and found that its ability to produce prodigiosin by fermentation increased by 21.74% compared with that of parent strain at pH4.0. These results indicated that xrpA regulates tolerance to low pH by transcriptional regulation of acid stress response genes to maintain cell membrane function in S. marcescens.
Project description:Global amphibian declines and extinction events are currently occurring at an unprecedented rate. While various factors are influencing these declines, one factor that is readily identifiable is disease. Specifically, the fungal pathogen Batrachochytrium dendrobatidis is thought to play a major role in amphibian declines in tropical and neotropical regions of the globe. While the effects of this chytrid fungus have been shown to be devastating, certain individuals and relict populations have shown resistance. This resistance has been attributed in part to the cutaneous microbiome. Many identified bacterial species that make up the microbiome have shown anti-B. dendrobatidis activity in vitro. One bacteria that is commonly associated as being a member of the amphibian microbiome across amphibian species and shows such anti-B. dendrobatidis activity is Serratia marcescens. Here, we look at transcriptomic shifts in gene expression of S. marcescens (high homology to strain WW4) in response to both live and heat-killed B. dendrobatidis.
Project description:Screening a library of 573 cyanobacteria extracts for inhibition of the quorum sensing regulated prodigiosin production of Serratia marcescens, an extract of the cyanobacterium Fischerella ambigua (Näg.) Gomont 108b was found to drastically increase the prodigiosin production. Bioactivity-guided isolation of the active compounds resulted in the two new natural products ambigol D and E along with the known ambigols A and C. Ambigol C treatment increased prodiginine production of Serratia sp. ATCC 39006 (S39006) by a factor of 10, while ambigols A and D were found to have antibiotic activity against this strain. RNA-Seq of S39006 treated with ambigol C and subsequent differential gene expression and functional enrichment analyses indicated a significant downregulation of genes associated with the translation machinery and fatty acid biosynthesis in Serratia, as well as increased expression of genes related to the uptake of l-proline. These results suggest that the ambigols increase the prodiginine production in S39006 not by activating the SmaIR quorum sensing system, but possibly by increasing the precursor supply of l-proline and malonyl-CoA.
Project description:In Aspergillus nidulans, expression of sulfur metabolism genes is activated by the MetR transcription factor containing a basic region and leucine zipper domain (bZIP). Here we identified and characterized MetZ, a new transcriptional regulator in Aspergillus nidulans and other Eurotiales. It contains a bZIP domain similar to the corresponding region in MetR and this similarity suggests that MetZ could potentially complement the MetR deficiency. The metR and metZ genes are interrupted by unusually long introns. Transcription of metZ, unlike that of metR, is controlled by the sulfur metabolite repression system (SMR) dependent on the MetR protein. Overexpression of metZ from a MetR-independent promoter in a metR strain, activates transcription of genes encoding sulfate permease, homocysteine synthase and methionine permease, partially complementing the phenotype of the metR mutation. Thus, MetZ appears to be a second transcription factor involved in regulation of sulfur metabolism genes.
Project description:metR (SPD_0588) mutant (TH9197) cannot grow in chemically defined medium (CDM) and had significant growth defect in CDM with 1 µg/ml methionine. With the increasing concentration of mehtionine, the growth of TH9197 was restored and the growth of metR mutant is as same as D39 WT (TH4306) when the concentration of methionine reaches 50 µg/ml. SPD_0588 is a putative transcriptional regulator and is homologous to MetR in Streptococcus mutans (78.9 % amino acid identity), which regulates both methionine synthesis and transport. We compared the transcriptome of WT and metR mutant in CDM with 1 µg/ml methionine by RNA-seq to see if MetR regulates the metabolism of methionine in Streptococcus pneumoniae D39. We compared the transcriptome of WT and metR mutant in CDM with 50 µg/ml methionine by RNA-seq to see if the regulation of MetR is methionine concentration-dependdent.We also want to identify the regulon of SPD_0588 in the two RNA-seq.