f nucleatum strain atcc 23726 genome  (ATCC)


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    ATCC f nucleatum strain atcc 23726 genome
    F Nucleatum Strain Atcc 23726 Genome, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    f nucleatum strain atcc 23726 genome  (ATCC)


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    ATCC f nucleatum strain atcc 23726 genome
    (A) Schematic diagram of assembling and working mode of the xylose-inducible system developed in this study to regulate reporter gene expression. This system was constructed by combining xylose repressor gene xylR , its operator sequence xylO , and the strong constitutive promoter P xylAB , the divergent promoter to drive the expression of Luciola Red luciferase ( luc ) gene reporter. The XylR is expressed constitutively from P xylR . It binds to the XylR operator (schematically depicted as a black) and blocks luc expression from the P xylAB promoter in the absence of xylose. XylR is released from the operator with xylose, initiating the luc expression. (B) The intergenic promoter region of the divergently transcribed xylR and luc in this xylose-inducible system. The translation initiation codon and the ribosomal bind site (RBS) are in bold red. The -10 and -35 sequences are shown as underlined sequences. The xylO element is highlighted in the box with dashed lines. Palindromic sequences in xylO are indicated by arrows. (C) Schematic diagram of assembling and working mode of the xylose and riboswitch-based dual inducible system developed in this study. The dual control system contains xylR , xylO , P xylAB, and a theophylline-responsive synthetical riboswitch E unit. The riboswitch is inserted to replace the RBS in the xylose-inducible system (A) . The reporter luc gene expression is initially repressed by XylR and is only activated in the presence of xylose. In the latter case, the luc gene is transcribed, but the mRNA is not translated in the absence of the riboswitch signal theophylline (RBS is hidden in a step-loop structure in riboswitch). In the presence of theophylline, the riboswitch changes its conformation and exposes the RBS so the ribosome can bind, resulting in synthesizing the luciferase protein. (D) The intergenic promoter region of the divergently transcribed xylR and luc in this dual inducible system. The linker region (underlined) and riboswitch E sequence were positioned as indicated in Figure (C). (E) Genetic maps of the xylose and riboswitch-based dual inducible expression reporter plasmid pBCG06 and the xylose-inducible alone system expression plasmid pBCG06a. Both plasmids are derivatives of E. coli-F. nucleatum shuttle vector pCWU6. Feature depicted: The xylR-PxylAB is from the vector pXCH, a xylose-inducible gene expression vector for Clostridium perfringens ; luc encodes a Luciola Red luciferase protein that is codon optimized for expression in low-GC bacteria. oriFn and repA , the replication region; oriEc , replication region of the E. coli plasmid pBR322; catP , the chloramphenicol acetyltransferase gene, conferring resistance to thiamphenicol in F. nucleatum or chloramphenicol in E. coli ; KanR is kanamycin resistance cassette from pJRD215. (F) Tunable induction of luciferase protein from P xylAB and the riboswitch control with different inducer(s) concentrations. The overnight cultures of F. nucleatum strain <t>ATCC</t> <t>23726</t> carrying pCWU6 (empty vector, EV), pBCG06, or pBCG06a were diluted 1:20 into TSPC. When cultures grew to OD600 ∼0.65, the inducers were added with various concentrations, as indicated. A tested culture received 20 mM glucose to assess its impact on the inducible system. After 2 hours of further incubation, 100 µl aliquots of each culture were taken out for luciferase assay. Luciferase activity (RLU) was normalized with cell density (OD 600 ). Each culture’s mean RLU/OD 600 was indicated above the corresponding column. The inserted graph showed the OD 600 value of the cultures when the luciferase assay was performed. Data represent the means of standard deviations of the results of triplicate biological experiments.
    F Nucleatum Strain Atcc 23726 Genome, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 86 stars, based on 1 article reviews
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    f nucleatum strain atcc 23726 genome - by Bioz Stars, 2024-05
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    1) Product Images from "Development of a Xylose-Inducible Promoter and Riboswitch Combination System for Manipulating Gene Expression in Fusobacterium nucleatum"

    Article Title: Development of a Xylose-Inducible Promoter and Riboswitch Combination System for Manipulating Gene Expression in Fusobacterium nucleatum

    Journal: bioRxiv

    doi: 10.1101/2023.04.24.538132

    (A) Schematic diagram of assembling and working mode of the xylose-inducible system developed in this study to regulate reporter gene expression. This system was constructed by combining xylose repressor gene xylR , its operator sequence xylO , and the strong constitutive promoter P xylAB , the divergent promoter to drive the expression of Luciola Red luciferase ( luc ) gene reporter. The XylR is expressed constitutively from P xylR . It binds to the XylR operator (schematically depicted as a black) and blocks luc expression from the P xylAB promoter in the absence of xylose. XylR is released from the operator with xylose, initiating the luc expression. (B) The intergenic promoter region of the divergently transcribed xylR and luc in this xylose-inducible system. The translation initiation codon and the ribosomal bind site (RBS) are in bold red. The -10 and -35 sequences are shown as underlined sequences. The xylO element is highlighted in the box with dashed lines. Palindromic sequences in xylO are indicated by arrows. (C) Schematic diagram of assembling and working mode of the xylose and riboswitch-based dual inducible system developed in this study. The dual control system contains xylR , xylO , P xylAB, and a theophylline-responsive synthetical riboswitch E unit. The riboswitch is inserted to replace the RBS in the xylose-inducible system (A) . The reporter luc gene expression is initially repressed by XylR and is only activated in the presence of xylose. In the latter case, the luc gene is transcribed, but the mRNA is not translated in the absence of the riboswitch signal theophylline (RBS is hidden in a step-loop structure in riboswitch). In the presence of theophylline, the riboswitch changes its conformation and exposes the RBS so the ribosome can bind, resulting in synthesizing the luciferase protein. (D) The intergenic promoter region of the divergently transcribed xylR and luc in this dual inducible system. The linker region (underlined) and riboswitch E sequence were positioned as indicated in Figure (C). (E) Genetic maps of the xylose and riboswitch-based dual inducible expression reporter plasmid pBCG06 and the xylose-inducible alone system expression plasmid pBCG06a. Both plasmids are derivatives of E. coli-F. nucleatum shuttle vector pCWU6. Feature depicted: The xylR-PxylAB is from the vector pXCH, a xylose-inducible gene expression vector for Clostridium perfringens ; luc encodes a Luciola Red luciferase protein that is codon optimized for expression in low-GC bacteria. oriFn and repA , the replication region; oriEc , replication region of the E. coli plasmid pBR322; catP , the chloramphenicol acetyltransferase gene, conferring resistance to thiamphenicol in F. nucleatum or chloramphenicol in E. coli ; KanR is kanamycin resistance cassette from pJRD215. (F) Tunable induction of luciferase protein from P xylAB and the riboswitch control with different inducer(s) concentrations. The overnight cultures of F. nucleatum strain ATCC 23726 carrying pCWU6 (empty vector, EV), pBCG06, or pBCG06a were diluted 1:20 into TSPC. When cultures grew to OD600 ∼0.65, the inducers were added with various concentrations, as indicated. A tested culture received 20 mM glucose to assess its impact on the inducible system. After 2 hours of further incubation, 100 µl aliquots of each culture were taken out for luciferase assay. Luciferase activity (RLU) was normalized with cell density (OD 600 ). Each culture’s mean RLU/OD 600 was indicated above the corresponding column. The inserted graph showed the OD 600 value of the cultures when the luciferase assay was performed. Data represent the means of standard deviations of the results of triplicate biological experiments.
    Figure Legend Snippet: (A) Schematic diagram of assembling and working mode of the xylose-inducible system developed in this study to regulate reporter gene expression. This system was constructed by combining xylose repressor gene xylR , its operator sequence xylO , and the strong constitutive promoter P xylAB , the divergent promoter to drive the expression of Luciola Red luciferase ( luc ) gene reporter. The XylR is expressed constitutively from P xylR . It binds to the XylR operator (schematically depicted as a black) and blocks luc expression from the P xylAB promoter in the absence of xylose. XylR is released from the operator with xylose, initiating the luc expression. (B) The intergenic promoter region of the divergently transcribed xylR and luc in this xylose-inducible system. The translation initiation codon and the ribosomal bind site (RBS) are in bold red. The -10 and -35 sequences are shown as underlined sequences. The xylO element is highlighted in the box with dashed lines. Palindromic sequences in xylO are indicated by arrows. (C) Schematic diagram of assembling and working mode of the xylose and riboswitch-based dual inducible system developed in this study. The dual control system contains xylR , xylO , P xylAB, and a theophylline-responsive synthetical riboswitch E unit. The riboswitch is inserted to replace the RBS in the xylose-inducible system (A) . The reporter luc gene expression is initially repressed by XylR and is only activated in the presence of xylose. In the latter case, the luc gene is transcribed, but the mRNA is not translated in the absence of the riboswitch signal theophylline (RBS is hidden in a step-loop structure in riboswitch). In the presence of theophylline, the riboswitch changes its conformation and exposes the RBS so the ribosome can bind, resulting in synthesizing the luciferase protein. (D) The intergenic promoter region of the divergently transcribed xylR and luc in this dual inducible system. The linker region (underlined) and riboswitch E sequence were positioned as indicated in Figure (C). (E) Genetic maps of the xylose and riboswitch-based dual inducible expression reporter plasmid pBCG06 and the xylose-inducible alone system expression plasmid pBCG06a. Both plasmids are derivatives of E. coli-F. nucleatum shuttle vector pCWU6. Feature depicted: The xylR-PxylAB is from the vector pXCH, a xylose-inducible gene expression vector for Clostridium perfringens ; luc encodes a Luciola Red luciferase protein that is codon optimized for expression in low-GC bacteria. oriFn and repA , the replication region; oriEc , replication region of the E. coli plasmid pBR322; catP , the chloramphenicol acetyltransferase gene, conferring resistance to thiamphenicol in F. nucleatum or chloramphenicol in E. coli ; KanR is kanamycin resistance cassette from pJRD215. (F) Tunable induction of luciferase protein from P xylAB and the riboswitch control with different inducer(s) concentrations. The overnight cultures of F. nucleatum strain ATCC 23726 carrying pCWU6 (empty vector, EV), pBCG06, or pBCG06a were diluted 1:20 into TSPC. When cultures grew to OD600 ∼0.65, the inducers were added with various concentrations, as indicated. A tested culture received 20 mM glucose to assess its impact on the inducible system. After 2 hours of further incubation, 100 µl aliquots of each culture were taken out for luciferase assay. Luciferase activity (RLU) was normalized with cell density (OD 600 ). Each culture’s mean RLU/OD 600 was indicated above the corresponding column. The inserted graph showed the OD 600 value of the cultures when the luciferase assay was performed. Data represent the means of standard deviations of the results of triplicate biological experiments.

    Techniques Used: Expressing, Construct, Sequencing, Luciferase, Plasmid Preparation, Incubation, Activity Assay

    (A) The effect of inducer(s) on the growth of conditional lepB mutants in the liquid media. F. nucleatum strains (wild-type: strain ATCC 23726 harboring an empty vector pCWU6 (red triangle); lepB conditional mutant: Δ lepB pAN01) were grown statically under anaerobic conditions on TSPC media with various doses of inducers (xylose and theophylline). Turbidity was monitored at an optical density of 600 nm. Data represented the means of standard deviation of the results of triplicate biological experiments. (B) The effects of inducer(s) on the growth of conditional lepB mutants in solid media. Ten-fold serial dilutions of overnight cultures of the wild-type 23726 and the conditional Δ lepB strains were spotted on agar plates with and without inducer(s) xylose and theophylline and cultivated in an anaerobic chamber at 37°C. Cell growth was recorded after three days of incubation. This experiment was repeated three times, and one representative experiment was shown. (C) LepB depletion creates short filaments and alters outer membrane structures. The lepB-depleted and lepB-inducing cells were immobilized on carbon-coated nickel grids and stained with 0.1% uranyl acetate before viewing with a transmission electron microscope. Enlarged areas of the top panels are shown in the bottom panels. Bars, 1 µm. (D) The altered cell surface of conditional lepB mutant was revealed by scanning electron microscopy. The cells from lepB depletion or lepB induction were immobilized on a silicon wafer and fixed with 2.5% glutaraldehyde before viewing by a scanning electron microscope. Enlarge areas of the top panel are shown in the below panels. Bars, 1 µm. (E) Comparison of lepB conditional mutant strain umorphology. Cells were grown under two distinct conditions with inducers (0.1% xylose/0.2 mM theophylline and 0.5% xylose/1.0 mM theophylline). Microscopic examination was conducted after cells reached the stationary phase to assess morphological differences. (F) The quantitative coaggregation assay was performed by mixing wild-type F. nucleatum , radD , and lepB mutants with an equal number of A. oris cells in the coaggregation buffer (CAB). The mixture was allowed to aggregate for 20 mins before measuring the OD 600 absorbance. Measurements were taken both before and after the aggregation process. The presented data represent the average of three independent experiments. Statistical analysis was performed using Student’s t -test with GraphPad Prism software.
    Figure Legend Snippet: (A) The effect of inducer(s) on the growth of conditional lepB mutants in the liquid media. F. nucleatum strains (wild-type: strain ATCC 23726 harboring an empty vector pCWU6 (red triangle); lepB conditional mutant: Δ lepB pAN01) were grown statically under anaerobic conditions on TSPC media with various doses of inducers (xylose and theophylline). Turbidity was monitored at an optical density of 600 nm. Data represented the means of standard deviation of the results of triplicate biological experiments. (B) The effects of inducer(s) on the growth of conditional lepB mutants in solid media. Ten-fold serial dilutions of overnight cultures of the wild-type 23726 and the conditional Δ lepB strains were spotted on agar plates with and without inducer(s) xylose and theophylline and cultivated in an anaerobic chamber at 37°C. Cell growth was recorded after three days of incubation. This experiment was repeated three times, and one representative experiment was shown. (C) LepB depletion creates short filaments and alters outer membrane structures. The lepB-depleted and lepB-inducing cells were immobilized on carbon-coated nickel grids and stained with 0.1% uranyl acetate before viewing with a transmission electron microscope. Enlarged areas of the top panels are shown in the bottom panels. Bars, 1 µm. (D) The altered cell surface of conditional lepB mutant was revealed by scanning electron microscopy. The cells from lepB depletion or lepB induction were immobilized on a silicon wafer and fixed with 2.5% glutaraldehyde before viewing by a scanning electron microscope. Enlarge areas of the top panel are shown in the below panels. Bars, 1 µm. (E) Comparison of lepB conditional mutant strain umorphology. Cells were grown under two distinct conditions with inducers (0.1% xylose/0.2 mM theophylline and 0.5% xylose/1.0 mM theophylline). Microscopic examination was conducted after cells reached the stationary phase to assess morphological differences. (F) The quantitative coaggregation assay was performed by mixing wild-type F. nucleatum , radD , and lepB mutants with an equal number of A. oris cells in the coaggregation buffer (CAB). The mixture was allowed to aggregate for 20 mins before measuring the OD 600 absorbance. Measurements were taken both before and after the aggregation process. The presented data represent the average of three independent experiments. Statistical analysis was performed using Student’s t -test with GraphPad Prism software.

    Techniques Used: Plasmid Preparation, Mutagenesis, Standard Deviation, Incubation, Staining, Transmission Assay, Microscopy, Electron Microscopy, Software

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    ATCC f nucleatum strain atcc 23726 genome
    F Nucleatum Strain Atcc 23726 Genome, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/f nucleatum strain atcc 23726 genome/product/ATCC
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    f nucleatum strain atcc 23726 genome - by Bioz Stars, 2024-05
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