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Biotechnology Information s. mutans ua159 genome annotation

Effect of smu_1361c on the growth, biofilm formation, and oxidative tolerance of S. mutans. (A) Growth characteristics of <t>UA159,</t> UA159/pDL278, UA159/pDL278-1361c, and UA159 Δ1361c. Bacteria were grown aerobically at 37°C, and OD600nm was monitored at 30-min intervals for 12 h. (B) The biofilm biomass was determined using crystal violet staining assay when bacteria were cultured in BHIS under aerobic conditions for 6, 12, and 24 h, respectively. The exponential cultures of UA159, UA159/pDL278, UA159/pDL278-1361c, and UA159 Δ1361c were exposed to oxidative stress with 0.2% (66.05 mM) hydrogen peroxide for 0, 20, 40, and 60 min, respectively, which were serially diluted and cultured on the BHI agar plates. The representative pictures of the hydrogen peroxide challenge are shown (C), colony-forming units were counted, and percent survivals of different strains were calculated relative to the control sample UA159 (D). The plates were overlaid with soft agar containing UA159, UA159/pDL278, UA159/pDL278-1361c, and UA159 Δ1361c and challenged with filter discs soaked with 0, 20, 40, and 60 mM hydrogen peroxide. The representative plates are shown to demonstrate the differences in inhibition zones (E), which are measured at three different positions and averaged to serve as a single data point, and the ratio of inhibition zone to disc diameter was calculated (F). Each experiment was repeated at least thrice. Results are presented as mean ± SD (*P < 0.05 or ***P < 0.001).

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Article Title: SMU_1361c regulates the oxidative stress response of Streptococcus mutans

Journal: Applied and Environmental Microbiology

doi: 10.1128/aem.01871-23

Effect of smu_1361c on the growth, biofilm formation, and oxidative tolerance of S. mutans. (A) Growth characteristics of UA159, UA159/pDL278, UA159/pDL278-1361c, and UA159 Δ1361c. Bacteria were grown aerobically at 37°C, and OD600nm was monitored at 30-min intervals for 12 h. (B) The biofilm biomass was determined using crystal violet staining assay when bacteria were cultured in BHIS under aerobic conditions for 6, 12, and 24 h, respectively. The exponential cultures of UA159, UA159/pDL278, UA159/pDL278-1361c, and UA159 Δ1361c were exposed to oxidative stress with 0.2% (66.05 mM) hydrogen peroxide for 0, 20, 40, and 60 min, respectively, which were serially diluted and cultured on the BHI agar plates. The representative pictures of the hydrogen peroxide challenge are shown (C), colony-forming units were counted, and percent survivals of different strains were calculated relative to the control sample UA159 (D). The plates were overlaid with soft agar containing UA159, UA159/pDL278, UA159/pDL278-1361c, and UA159 Δ1361c and challenged with filter discs soaked with 0, 20, 40, and 60 mM hydrogen peroxide. The representative plates are shown to demonstrate the differences in inhibition zones (E), which are measured at three different positions and averaged to serve as a single data point, and the ratio of inhibition zone to disc diameter was calculated (F). Each experiment was repeated at least thrice. Results are presented as mean ± SD (*P < 0.05 or ***P < 0.001).

Figure Legend Snippet: Effect of smu_1361c on the growth, biofilm formation, and oxidative tolerance of S. mutans. (A) Growth characteristics of UA159, UA159/pDL278, UA159/pDL278-1361c, and UA159 Δ1361c. Bacteria were grown aerobically at 37°C, and OD600nm was monitored at 30-min intervals for 12 h. (B) The biofilm biomass was determined using crystal violet staining assay when bacteria were cultured in BHIS under aerobic conditions for 6, 12, and 24 h, respectively. The exponential cultures of UA159, UA159/pDL278, UA159/pDL278-1361c, and UA159 Δ1361c were exposed to oxidative stress with 0.2% (66.05 mM) hydrogen peroxide for 0, 20, 40, and 60 min, respectively, which were serially diluted and cultured on the BHI agar plates. The representative pictures of the hydrogen peroxide challenge are shown (C), colony-forming units were counted, and percent survivals of different strains were calculated relative to the control sample UA159 (D). The plates were overlaid with soft agar containing UA159, UA159/pDL278, UA159/pDL278-1361c, and UA159 Δ1361c and challenged with filter discs soaked with 0, 20, 40, and 60 mM hydrogen peroxide. The representative plates are shown to demonstrate the differences in inhibition zones (E), which are measured at three different positions and averaged to serve as a single data point, and the ratio of inhibition zone to disc diameter was calculated (F). Each experiment was repeated at least thrice. Results are presented as mean ± SD (*P < 0.05 or ***P < 0.001).

Techniques Used: Bacteria, Staining, Cell Culture, Control, Inhibition

Effect of smu_1361c on interspecies competition within the three-species biofilms. S. mutans and its derivatives, S. gordonii, and S. sanguinis were simultaneously cultured in fresh BHIS (A and B) or supplemented with 3 mg/mL catalase (C and D) under aerobic conditions for 24 h and labeled by species-specific FISH probes. The three-dimensional visualization of three-species biofilms with S. mutans (green), S. gordonii (blue), and S. sanguinis (red) was captured using the confocal laser scanning microscopy at 60× magnification and reconstructed using the IMARIS 7.0.0 (A and C). The ratio of S. mutans/S. gordonii/S. sanguinis was quantified by the coverage area of each species with Image Pro Plus 6.0 (B and D). The representative images are shown from at least five randomly selected positions of each sample. Results are presented as mean ± SD (*P < 0.05 or ****P < 0.0001).

Figure Legend Snippet: Effect of smu_1361c on interspecies competition within the three-species biofilms. S. mutans and its derivatives, S. gordonii, and S. sanguinis were simultaneously cultured in fresh BHIS (A and B) or supplemented with 3 mg/mL catalase (C and D) under aerobic conditions for 24 h and labeled by species-specific FISH probes. The three-dimensional visualization of three-species biofilms with S. mutans (green), S. gordonii (blue), and S. sanguinis (red) was captured using the confocal laser scanning microscopy at 60× magnification and reconstructed using the IMARIS 7.0.0 (A and C). The ratio of S. mutans/S. gordonii/S. sanguinis was quantified by the coverage area of each species with Image Pro Plus 6.0 (B and D). The representative images are shown from at least five randomly selected positions of each sample. Results are presented as mean ± SD (*P < 0.05 or ****P < 0.0001).

Techniques Used: Cell Culture, Labeling, Confocal Laser Scanning Microscopy

Transcriptomics analysis of UA159/pDL278 and UA159/pDL278-1361c strains. (A) Functional classification of differentially expressed genes (DEGs) was performed based on the clusters of orthologous groups type. (B) The volcano plot illustrates DEGs between UA159/pDL278 and UA159/pDL278-1361c. Functional annotation and enrichment analysis of DEGs were performed in the Kyoto Encyclopedia of Genes and Genomes (KEGG, C) and Gene Ontology (GO, D) databases. Downregulated genes are shown in green. (E) Genetic organization of gene clusters that were significantly downregulated in UA159/pDL278-1361c.

Figure Legend Snippet: Transcriptomics analysis of UA159/pDL278 and UA159/pDL278-1361c strains. (A) Functional classification of differentially expressed genes (DEGs) was performed based on the clusters of orthologous groups type. (B) The volcano plot illustrates DEGs between UA159/pDL278 and UA159/pDL278-1361c. Functional annotation and enrichment analysis of DEGs were performed in the Kyoto Encyclopedia of Genes and Genomes (KEGG, C) and Gene Ontology (GO, D) databases. Downregulated genes are shown in green. (E) Genetic organization of gene clusters that were significantly downregulated in UA159/pDL278-1361c.

Techniques Used: Functional Assay

Figure Legend Snippet: Bacterial strains and plasmids used in this study

Techniques Used: Expressing, Plasmid Preparation, Sequencing, Over Expression

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Journal: Applied and Environmental Microbiology

Article Title: SMU_1361c regulates the oxidative stress response of Streptococcus mutans

doi: 10.1128/aem.01871-23

Figure Lengend Snippet: Effect of smu_1361c on the growth, biofilm formation, and oxidative tolerance of S. mutans. (A) Growth characteristics of UA159, UA159/pDL278, UA159/pDL278-1361c, and UA159 Δ1361c. Bacteria were grown aerobically at 37°C, and OD600nm was monitored at 30-min intervals for 12 h. (B) The biofilm biomass was determined using crystal violet staining assay when bacteria were cultured in BHIS under aerobic conditions for 6, 12, and 24 h, respectively. The exponential cultures of UA159, UA159/pDL278, UA159/pDL278-1361c, and UA159 Δ1361c were exposed to oxidative stress with 0.2% (66.05 mM) hydrogen peroxide for 0, 20, 40, and 60 min, respectively, which were serially diluted and cultured on the BHI agar plates. The representative pictures of the hydrogen peroxide challenge are shown (C), colony-forming units were counted, and percent survivals of different strains were calculated relative to the control sample UA159 (D). The plates were overlaid with soft agar containing UA159, UA159/pDL278, UA159/pDL278-1361c, and UA159 Δ1361c and challenged with filter discs soaked with 0, 20, 40, and 60 mM hydrogen peroxide. The representative plates are shown to demonstrate the differences in inhibition zones (E), which are measured at three different positions and averaged to serve as a single data point, and the ratio of inhibition zone to disc diameter was calculated (F). Each experiment was repeated at least thrice. Results are presented as mean ± SD (*P < 0.05 or ***P < 0.001).

Article Snippet: Based on the S. mutans UA159 genome annotation from the National Center for Biotechnology Information, most of the significantly downregulated genes were mainly associated with energy production and conversion and carbohydrate transport and metabolism, except for some unannotated genes ( ).

Techniques: Bacteria, Staining, Cell Culture, Control, Inhibition

Journal: Applied and Environmental Microbiology

Article Title: SMU_1361c regulates the oxidative stress response of Streptococcus mutans

doi: 10.1128/aem.01871-23

Figure Lengend Snippet: Effect of smu_1361c on interspecies competition within the three-species biofilms. S. mutans and its derivatives, S. gordonii, and S. sanguinis were simultaneously cultured in fresh BHIS (A and B) or supplemented with 3 mg/mL catalase (C and D) under aerobic conditions for 24 h and labeled by species-specific FISH probes. The three-dimensional visualization of three-species biofilms with S. mutans (green), S. gordonii (blue), and S. sanguinis (red) was captured using the confocal laser scanning microscopy at 60× magnification and reconstructed using the IMARIS 7.0.0 (A and C). The ratio of S. mutans/S. gordonii/S. sanguinis was quantified by the coverage area of each species with Image Pro Plus 6.0 (B and D). The representative images are shown from at least five randomly selected positions of each sample. Results are presented as mean ± SD (*P < 0.05 or ****P < 0.0001).

Techniques: Cell Culture, Labeling, Confocal Laser Scanning Microscopy

Journal: Applied and Environmental Microbiology

Article Title: SMU_1361c regulates the oxidative stress response of Streptococcus mutans

doi: 10.1128/aem.01871-23

Figure Lengend Snippet: Transcriptomics analysis of UA159/pDL278 and UA159/pDL278-1361c strains. (A) Functional classification of differentially expressed genes (DEGs) was performed based on the clusters of orthologous groups type. (B) The volcano plot illustrates DEGs between UA159/pDL278 and UA159/pDL278-1361c. Functional annotation and enrichment analysis of DEGs were performed in the Kyoto Encyclopedia of Genes and Genomes (KEGG, C) and Gene Ontology (GO, D) databases. Downregulated genes are shown in green. (E) Genetic organization of gene clusters that were significantly downregulated in UA159/pDL278-1361c.

Techniques: Functional Assay

Journal: Applied and Environmental Microbiology

Article Title: SMU_1361c regulates the oxidative stress response of Streptococcus mutans

doi: 10.1128/aem.01871-23

Figure Lengend Snippet: Bacterial strains and plasmids used in this study

Techniques: Expressing, Plasmid Preparation, Sequencing, Over Expression

Deletion of stsR decreases S. mutans biofilm formation at early stage. S. mutans was cultured in BM supplemented with 1% sucrose for 6, 12, 24, and 48 h. The biofilm biomass was determined by CV staining method. Data from three biological replicates were averaged, and the statistical significance between the stsR mutant, wild-type, and complement strain was determined by Student’s t -test. Error bars represent standard deviations based on results from at least three biological replicates. ∗∗ Indicates a significance of P < 0.01.

Journal: Frontiers in Microbiology

Article Title: A GntR Family Transcription Factor in Streptococcus mutans Regulates Biofilm Formation and Expression of Multiple Sugar Transporter Genes

doi: 10.3389/fmicb.2018.03224

Figure Lengend Snippet: Deletion of stsR decreases S. mutans biofilm formation at early stage. S. mutans was cultured in BM supplemented with 1% sucrose for 6, 12, 24, and 48 h. The biofilm biomass was determined by CV staining method. Data from three biological replicates were averaged, and the statistical significance between the stsR mutant, wild-type, and complement strain was determined by Student’s t -test. Error bars represent standard deviations based on results from at least three biological replicates. ∗∗ Indicates a significance of P < 0.01.

Article Snippet: According to the National Center for Biotechnology Information (NCBI) S. mutans genome annotation, the function of the majority of the differentially expressed genes (DEGs) are unknown.

Techniques: Cell Culture, Staining, Mutagenesis

Determination of growth curves of S. mutans UA159, S. mutans Δ stsR , and Δ stsR/pDL278-stsR . S. mutans UA159, S. mutans Δ stsR , and Δ stsR/pDL278-stsR strains were cultivated in BHI to mid-exponential phase and then diluted into (A) fresh BHI broth or (B) TV base medium supplemented with either 10 mM (limiting) or 100 mM (excess) sucrose, glucose, or lactose. Growth curves were monitored with a Multiskan Spectrum (Thermo, Multiskan Go, United States), and the OD 600 was measured in 1 h intervals. (A) For CFU counts, after diluted, the bacteria were cultured at 37°C for 1, 4, 8, and 12 h. Then bacterial suspension was serially diluted in BHI and plated on BHI agar plates. CFU values were calculated after the plates were incubated anaerobically at 37°C for 48 h. Error bars represent standard deviations based on results from at least three biological replicates.

Journal: Frontiers in Microbiology

Article Title: A GntR Family Transcription Factor in Streptococcus mutans Regulates Biofilm Formation and Expression of Multiple Sugar Transporter Genes

doi: 10.3389/fmicb.2018.03224

Figure Lengend Snippet: Determination of growth curves of S. mutans UA159, S. mutans Δ stsR , and Δ stsR/pDL278-stsR . S. mutans UA159, S. mutans Δ stsR , and Δ stsR/pDL278-stsR strains were cultivated in BHI to mid-exponential phase and then diluted into (A) fresh BHI broth or (B) TV base medium supplemented with either 10 mM (limiting) or 100 mM (excess) sucrose, glucose, or lactose. Growth curves were monitored with a Multiskan Spectrum (Thermo, Multiskan Go, United States), and the OD 600 was measured in 1 h intervals. (A) For CFU counts, after diluted, the bacteria were cultured at 37°C for 1, 4, 8, and 12 h. Then bacterial suspension was serially diluted in BHI and plated on BHI agar plates. CFU values were calculated after the plates were incubated anaerobically at 37°C for 48 h. Error bars represent standard deviations based on results from at least three biological replicates.

Techniques: Bacteria, Cell Culture, Suspension, Incubation

Deletion of stsR decreases the amount of glucans in S. mutans biofilm. The amounts of (A) water insoluble glucans and (B) water soluble glucans in the biofilms of S. mutans UA159, S. mutans Δ stsR , and complement strain were quantified using the phenol-sulfuric acid method and calculated according to the standard curve. Error bars represent standard deviations based on results from at least three biological replicates. ∗∗ Indicates a significance of P < 0.01.

Journal: Frontiers in Microbiology

Article Title: A GntR Family Transcription Factor in Streptococcus mutans Regulates Biofilm Formation and Expression of Multiple Sugar Transporter Genes

doi: 10.3389/fmicb.2018.03224

Figure Lengend Snippet: Deletion of stsR decreases the amount of glucans in S. mutans biofilm. The amounts of (A) water insoluble glucans and (B) water soluble glucans in the biofilms of S. mutans UA159, S. mutans Δ stsR , and complement strain were quantified using the phenol-sulfuric acid method and calculated according to the standard curve. Error bars represent standard deviations based on results from at least three biological replicates. ∗∗ Indicates a significance of P < 0.01.

Techniques:

Scanning electron microscopy analysis reveals altered biofilm morphology and decreased biofilm extracellular matrix of S. mutans Δ stsR . Biofilms formed by S. mutans UA159, S. mutans Δ stsR , and complement strain were grown for 6 h and then scanned by scanning electron microscopy (SEM) under (A) 1000× magnification, (B) 5000× magnification, and (C) 20000× magnification.

Journal: Frontiers in Microbiology

Article Title: A GntR Family Transcription Factor in Streptococcus mutans Regulates Biofilm Formation and Expression of Multiple Sugar Transporter Genes

doi: 10.3389/fmicb.2018.03224

Figure Lengend Snippet: Scanning electron microscopy analysis reveals altered biofilm morphology and decreased biofilm extracellular matrix of S. mutans Δ stsR . Biofilms formed by S. mutans UA159, S. mutans Δ stsR , and complement strain were grown for 6 h and then scanned by scanning electron microscopy (SEM) under (A) 1000× magnification, (B) 5000× magnification, and (C) 20000× magnification.

Techniques: Electron Microscopy

Biofilm structure and EPS distribution of S. mutans strains observed by confocal microscopy. (A) Double-labeling of 6 h S. mutans biofilms. Green indicates bacteria (SYTO 9), and red indicates EPS (Alexa Fluor 647). Images were taken at 60× magnification. The three-dimensional reconstruction of the biofilms and the quantification of EPS/bacteria biomass were performed with IMARIS 7.0.0. (B) The ratio of EPS to bacteria at different heights was quantified with COMSTAT. Results are the average of five randomly selected positions of each sample and are presented as mean ± standard deviation. (C–E) Quantification of S. mutans UA159 (C) , S. mutans Δ stsR (D) , and Δ stsR/pDL278-stsR (E) biofilms. EPS biomass was performed with COMSTAT at different heights. Results are the average of five randomly selected positions of each sample and are presented as mean ± standard deviation.

Journal: Frontiers in Microbiology

Article Title: A GntR Family Transcription Factor in Streptococcus mutans Regulates Biofilm Formation and Expression of Multiple Sugar Transporter Genes

doi: 10.3389/fmicb.2018.03224

Figure Lengend Snippet: Biofilm structure and EPS distribution of S. mutans strains observed by confocal microscopy. (A) Double-labeling of 6 h S. mutans biofilms. Green indicates bacteria (SYTO 9), and red indicates EPS (Alexa Fluor 647). Images were taken at 60× magnification. The three-dimensional reconstruction of the biofilms and the quantification of EPS/bacteria biomass were performed with IMARIS 7.0.0. (B) The ratio of EPS to bacteria at different heights was quantified with COMSTAT. Results are the average of five randomly selected positions of each sample and are presented as mean ± standard deviation. (C–E) Quantification of S. mutans UA159 (C) , S. mutans Δ stsR (D) , and Δ stsR/pDL278-stsR (E) biofilms. EPS biomass was performed with COMSTAT at different heights. Results are the average of five randomly selected positions of each sample and are presented as mean ± standard deviation.

Techniques: Confocal Microscopy, Labeling, Bacteria, Standard Deviation

Quantitative RT-PCR assays for the relative expression levels of gtfs and ftf genes in S. mutans UA159, S. mutans Δ stsR , and Δ stsR/pDL278-stsR . The experiments were carried out as described in Experimental procedures. All target genes were amplified using specific primers. Different gene expressions were normalized to the levels of 16S rRNA gene transcripts, and the folds of expression change were calculated.

Journal: Frontiers in Microbiology

Article Title: A GntR Family Transcription Factor in Streptococcus mutans Regulates Biofilm Formation and Expression of Multiple Sugar Transporter Genes

doi: 10.3389/fmicb.2018.03224

Figure Lengend Snippet: Quantitative RT-PCR assays for the relative expression levels of gtfs and ftf genes in S. mutans UA159, S. mutans Δ stsR , and Δ stsR/pDL278-stsR . The experiments were carried out as described in Experimental procedures. All target genes were amplified using specific primers. Different gene expressions were normalized to the levels of 16S rRNA gene transcripts, and the folds of expression change were calculated.

Techniques: Quantitative RT-PCR, Expressing, Amplification

The differentially expressed sugar transport system operons in S. mutans Δ stsR . The genetic organization of differentially expressed gene clusters that were associated with sugar transport systems in S. mutans Δ stsR . Upregulated genes are colored red, and downregulated genes are colored green.

Journal: Frontiers in Microbiology

Article Title: A GntR Family Transcription Factor in Streptococcus mutans Regulates Biofilm Formation and Expression of Multiple Sugar Transporter Genes

doi: 10.3389/fmicb.2018.03224

Figure Lengend Snippet: The differentially expressed sugar transport system operons in S. mutans Δ stsR . The genetic organization of differentially expressed gene clusters that were associated with sugar transport systems in S. mutans Δ stsR . Upregulated genes are colored red, and downregulated genes are colored green.

Techniques:

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