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ATCC strain atcc3624

Figure 1. Expression of brnQ genes during culture in TY cultures of <t>ATCC3624.</t> (A) RT-PCR analyses. The recA gene was used as a housekeeping gene, and a sample lacking reverse transcriptase (RT-) was run (top panel) in a PCR reaction with recA primers to show the absence of DNA contamination in these RNA samples; recA gene RT-PCR (2nd panel) showed that cDNA was present in all samples. Further RT-PCR analyses (bottom three panels) are shown using brnQ1 primers (for brnQ1 expres- sion), brnQ2 primers (for brnQ2 expression), or brnQ3 primers (for brnQ3 expression) for wild-type ATCC3624 cultures at different time points. The expected size of each PCR product is listed in Table 1. The sample with ATCC3624 DNA served as the positive control, and the sample without ATCC3624 DNA served as the negative control. DNA size markers are shown on the left of each gel. The results shown are representative of three repetitions. (B) A comparison of the RT-PCR band intensity ratio between the brnQ gene vs. the housekeeping recA gene is shown using the Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the 2 h result.

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1) Product Images from "BrnQ Branched-Chain Amino Acid Transporters Influence Toxin Production by, but Not Growth of, Clostridium perfringens Type A Strain ATCC3624."

Article Title: BrnQ Branched-Chain Amino Acid Transporters Influence Toxin Production by, but Not Growth of, Clostridium perfringens Type A Strain ATCC3624.

Journal: Toxins

doi: 10.3390/toxins17040187

Figure 1. Expression of brnQ genes during culture in TY cultures of ATCC3624. (A) RT-PCR analyses. The recA gene was used as a housekeeping gene, and a sample lacking reverse transcriptase (RT-) was run (top panel) in a PCR reaction with recA primers to show the absence of DNA contamination in these RNA samples; recA gene RT-PCR (2nd panel) showed that cDNA was present in all samples. Further RT-PCR analyses (bottom three panels) are shown using brnQ1 primers (for brnQ1 expres- sion), brnQ2 primers (for brnQ2 expression), or brnQ3 primers (for brnQ3 expression) for wild-type ATCC3624 cultures at different time points. The expected size of each PCR product is listed in Table 1. The sample with ATCC3624 DNA served as the positive control, and the sample without ATCC3624 DNA served as the negative control. DNA size markers are shown on the left of each gel. The results shown are representative of three repetitions. (B) A comparison of the RT-PCR band intensity ratio between the brnQ gene vs. the housekeeping recA gene is shown using the Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the 2 h result.

Figure Legend Snippet: Figure 1. Expression of brnQ genes during culture in TY cultures of ATCC3624. (A) RT-PCR analyses. The recA gene was used as a housekeeping gene, and a sample lacking reverse transcriptase (RT-) was run (top panel) in a PCR reaction with recA primers to show the absence of DNA contamination in these RNA samples; recA gene RT-PCR (2nd panel) showed that cDNA was present in all samples. Further RT-PCR analyses (bottom three panels) are shown using brnQ1 primers (for brnQ1 expres- sion), brnQ2 primers (for brnQ2 expression), or brnQ3 primers (for brnQ3 expression) for wild-type ATCC3624 cultures at different time points. The expected size of each PCR product is listed in Table 1. The sample with ATCC3624 DNA served as the positive control, and the sample without ATCC3624 DNA served as the negative control. DNA size markers are shown on the left of each gel. The results shown are representative of three repetitions. (B) A comparison of the RT-PCR band intensity ratio between the brnQ gene vs. the housekeeping recA gene is shown using the Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the 2 h result.

Techniques Used: Expressing, Reverse Transcription Polymerase Chain Reaction, Reverse Transcription, Positive Control, Negative Control, Comparison, Software

Figure 2. A comparison of post-inoculation changes in culture OD600, vegetative cell viability, and PLC or PFO production levels for ATCC3624 (WT), its brnQ1 null mutant (brnQ1KO), and the complementing strain (brnQ1c) when cultured in a TY medium. (A) Post-inoculation changes in the OD600 of TY cultures of these strains from 0 to 9 h at 37 ◦C. The result shown is representative of three repetitions. (B) OD600 of TY cultures at 24 h at 37 ◦C (top). The same samples were also used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. (C) Western blot analyses of PLC level in supernatants of 3, 7, and 24 h culture samples (top panel). The PLC blot was also repeated using 10 µL more of the sample from the same 3 h TY sample, with longer film exposure (bottom left blot), to allow for a better comparison of PLC production between the brnQ1 null mutant vs. wild-type ATCC3624 or the complementing strain. The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio is shown between 3 h samples (bottom right graph) using Image J software. The graph shows the mean values from three independent experiments. The error bars indicate the S.D. * p < 0.05 relative to wild type. Similar Image J analyses did not detect PLC production differences among these strains in 7 or 24 h samples (Figure S5A). (D) Western blot analyses of the PFO level in supernatants of 3, 7, and 24 h culture samples. The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. Image J analyses did not detect significant differences in PFO production among these strains at any time points (Figure S5A).

Figure Legend Snippet: Figure 2. A comparison of post-inoculation changes in culture OD600, vegetative cell viability, and PLC or PFO production levels for ATCC3624 (WT), its brnQ1 null mutant (brnQ1KO), and the complementing strain (brnQ1c) when cultured in a TY medium. (A) Post-inoculation changes in the OD600 of TY cultures of these strains from 0 to 9 h at 37 ◦C. The result shown is representative of three repetitions. (B) OD600 of TY cultures at 24 h at 37 ◦C (top). The same samples were also used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. (C) Western blot analyses of PLC level in supernatants of 3, 7, and 24 h culture samples (top panel). The PLC blot was also repeated using 10 µL more of the sample from the same 3 h TY sample, with longer film exposure (bottom left blot), to allow for a better comparison of PLC production between the brnQ1 null mutant vs. wild-type ATCC3624 or the complementing strain. The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio is shown between 3 h samples (bottom right graph) using Image J software. The graph shows the mean values from three independent experiments. The error bars indicate the S.D. * p < 0.05 relative to wild type. Similar Image J analyses did not detect PLC production differences among these strains in 7 or 24 h samples (Figure S5A). (D) Western blot analyses of the PFO level in supernatants of 3, 7, and 24 h culture samples. The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. Image J analyses did not detect significant differences in PFO production among these strains at any time points (Figure S5A).

Techniques Used: Comparison, Mutagenesis, Cell Culture, Western Blot, Software

Figure 3. A comparison of post-inoculation changes in culture OD600, vegetative cell viability, and PLC or PFO production levels for ATCC3624 (WT), its brnQ2 null mutant (brnQ2KO), and the complementing strain (brnQ2c) when cultured in TY medium. (A) Post-inoculation changes in the OD600 of TY cultures of these strains from 0 to 9 h at 37 ◦C. The result shown is representative of three repetitions. (B) The OD600 of TY cultures at 24 h at 37 ◦C (top). The same samples were used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. (C) Western blot analyses of PLC levels in supernatants of 3, 7, and 24 h culture samples (top panel). The PLC blot was also repeated using 10 µL more of the same 3 h TY samples, with longer film exposure (bottom left blot), to allow for a better comparison of PLC production between the brnQ2 null mutant vs. wild-type ATCC3624 or the complementing strain. The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio is shown between 3 h samples (bottom right graph) using Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect PLC production differences among the strains in 7 or 24 h samples (Figure S5B). (D) Western blot analyses of PFO levels in supernatants of 3, 7, and 24 h culture samples. The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown at left. Image J software did not detect significant differences in PFO production among the strains at any time points (Figure S5B).

Figure Legend Snippet: Figure 3. A comparison of post-inoculation changes in culture OD600, vegetative cell viability, and PLC or PFO production levels for ATCC3624 (WT), its brnQ2 null mutant (brnQ2KO), and the complementing strain (brnQ2c) when cultured in TY medium. (A) Post-inoculation changes in the OD600 of TY cultures of these strains from 0 to 9 h at 37 ◦C. The result shown is representative of three repetitions. (B) The OD600 of TY cultures at 24 h at 37 ◦C (top). The same samples were used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. (C) Western blot analyses of PLC levels in supernatants of 3, 7, and 24 h culture samples (top panel). The PLC blot was also repeated using 10 µL more of the same 3 h TY samples, with longer film exposure (bottom left blot), to allow for a better comparison of PLC production between the brnQ2 null mutant vs. wild-type ATCC3624 or the complementing strain. The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio is shown between 3 h samples (bottom right graph) using Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect PLC production differences among the strains in 7 or 24 h samples (Figure S5B). (D) Western blot analyses of PFO levels in supernatants of 3, 7, and 24 h culture samples. The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown at left. Image J software did not detect significant differences in PFO production among the strains at any time points (Figure S5B).

Techniques Used: Comparison, Mutagenesis, Cell Culture, Western Blot, Software

Figure 4. A comparison of the post-inoculation changes in culture OD600, vegetative cell viability, and PLC or PFO production levels for ATCC3624 (WT), its brnQ3 null mutant (brnQ3KO), and the complementing strain (brnQ3c) when cultured in a TY medium. (A) Post-inoculation changes in the OD600 of TY cultures from 0 to 9 h at 37 ◦C. The result shown is representative of three repetitions. (B) The OD600 of TY cultures at 24 h at 37 ◦C (top). The same samples were also used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. (C) Western blot analyses of PLC levels in supernatants of 3, 7, and 24 h culture samples (top panel). The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio is shown between 3 h samples (bottom graph) using Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect PLC production differences among these strains in 7 or 24 h samples (Figure S5C). (D) Western blot analyses of PFO levels in supernatants of 3, 7, and 24 h culture samples. The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. Image J analyses did not detect significant differences in PFO production among these strains at any time points (Figure S5C).

Figure Legend Snippet: Figure 4. A comparison of the post-inoculation changes in culture OD600, vegetative cell viability, and PLC or PFO production levels for ATCC3624 (WT), its brnQ3 null mutant (brnQ3KO), and the complementing strain (brnQ3c) when cultured in a TY medium. (A) Post-inoculation changes in the OD600 of TY cultures from 0 to 9 h at 37 ◦C. The result shown is representative of three repetitions. (B) The OD600 of TY cultures at 24 h at 37 ◦C (top). The same samples were also used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. (C) Western blot analyses of PLC levels in supernatants of 3, 7, and 24 h culture samples (top panel). The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio is shown between 3 h samples (bottom graph) using Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect PLC production differences among these strains in 7 or 24 h samples (Figure S5C). (D) Western blot analyses of PFO levels in supernatants of 3, 7, and 24 h culture samples. The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. Image J analyses did not detect significant differences in PFO production among these strains at any time points (Figure S5C).

Techniques Used: Comparison, Mutagenesis, Cell Culture, Western Blot, Software

Figure 5. A comparison of expression of the brnQ genes by wild-type ATCC3624, its isogenic brnQ null mutants, and complementing strains. ATCC3624, the single brnQ null mutants, or complementing strains were inoculated into TY broth and then incubated anaerobically for 2 h at 37 ◦C. Bacteria were collected and pelleted via centrifugation. Total RNA was extracted from the pellets, and cDNA was made for RT-qPCR analyses. Average CT values were normalized to the housekeeping recA gene, and the fold differences in expression were calculated using the comparative CT method (2−∆∆CT). The mean values from three independent experiments are shown. The error bars indicate the S.D. (A) RT- qPCR for the expression of brnQ2 or brnQ3 was performed using cDNA from the wild-type parent, the brnQ1 null mutant, and the complementing strain after their incubation in a TY medium for 2 h. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. (B) RT-qPCR for the expression of brnQ1 or brnQ3 was performed using cDNA from the wild type-parent, the brnQ2 null mutant, and the complementing strain after their incubation in TY medium for 2 h. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. (C) RT-qPCR for the expression of brnQ1 or brnQ2 was performed using cDNA from the wild-type parent, the brnQ3 null mutant, and the complementing strain after their incubation in TY medium for 2 h. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type.

Figure Legend Snippet: Figure 5. A comparison of expression of the brnQ genes by wild-type ATCC3624, its isogenic brnQ null mutants, and complementing strains. ATCC3624, the single brnQ null mutants, or complementing strains were inoculated into TY broth and then incubated anaerobically for 2 h at 37 ◦C. Bacteria were collected and pelleted via centrifugation. Total RNA was extracted from the pellets, and cDNA was made for RT-qPCR analyses. Average CT values were normalized to the housekeeping recA gene, and the fold differences in expression were calculated using the comparative CT method (2−∆∆CT). The mean values from three independent experiments are shown. The error bars indicate the S.D. (A) RT- qPCR for the expression of brnQ2 or brnQ3 was performed using cDNA from the wild-type parent, the brnQ1 null mutant, and the complementing strain after their incubation in a TY medium for 2 h. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. (B) RT-qPCR for the expression of brnQ1 or brnQ3 was performed using cDNA from the wild type-parent, the brnQ2 null mutant, and the complementing strain after their incubation in TY medium for 2 h. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. (C) RT-qPCR for the expression of brnQ1 or brnQ2 was performed using cDNA from the wild-type parent, the brnQ3 null mutant, and the complementing strain after their incubation in TY medium for 2 h. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type.

Techniques Used: Comparison, Expressing, Incubation, Bacteria, Centrifugation, Quantitative RT-PCR, Mutagenesis

Figure 7. A comparison of PLC or PFO production levels for wild-type ATCC3624, its double null mutants, and the complementing strain when cultured in a TY medium. Western blot analyses of PLC (A) and PFO (B) levels in supernatants of the wild type, double null mutants, and their complementing strains in the early-stage culture (3 h). The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. The right figure in panels A and B show the comparison of Western blot intensities determined by Image J. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type.

Figure Legend Snippet: Figure 7. A comparison of PLC or PFO production levels for wild-type ATCC3624, its double null mutants, and the complementing strain when cultured in a TY medium. Western blot analyses of PLC (A) and PFO (B) levels in supernatants of the wild type, double null mutants, and their complementing strains in the early-stage culture (3 h). The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. The right figure in panels A and B show the comparison of Western blot intensities determined by Image J. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type.

Techniques Used: Comparison, Cell Culture, Western Blot

Figure 8. A comparison of post-inoculation changes in culture OD600, vegetative cell viability, or PLC and PFO production levels for ATCC3624 (WT) vs. the triple null mutant (TKO) when cultured in a TY medium. (A) Post-inoculation changes in the OD600 of TY cultures incubated for up to 9 h at 37 ◦C. The results shown are representative of three repetitions. (B) The OD600 of TY cultures after 24 h of incubation at 37 ◦C (top). The same samples were used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. (C) Western blot analyses of PLC levels in the supernatants of panel A and B samples (3 h, 7 h and 24 h). The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio between 3 h samples (right graph) using Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect any PLC production differences among these strains in 7 or 24 h samples (Figure S10A). (D) Western blot analyses of PFO levels in the supernatants of panel A and B samples (3 h, 7 h and 24 h). The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. The right graph in panel D shows the comparison of the Western blot band intensity ratio between 3 h samples using the Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect any PFO production differences among these strains in 7 or 24 h samples (Figure S10B).

Figure Legend Snippet: Figure 8. A comparison of post-inoculation changes in culture OD600, vegetative cell viability, or PLC and PFO production levels for ATCC3624 (WT) vs. the triple null mutant (TKO) when cultured in a TY medium. (A) Post-inoculation changes in the OD600 of TY cultures incubated for up to 9 h at 37 ◦C. The results shown are representative of three repetitions. (B) The OD600 of TY cultures after 24 h of incubation at 37 ◦C (top). The same samples were used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. (C) Western blot analyses of PLC levels in the supernatants of panel A and B samples (3 h, 7 h and 24 h). The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio between 3 h samples (right graph) using Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect any PLC production differences among these strains in 7 or 24 h samples (Figure S10A). (D) Western blot analyses of PFO levels in the supernatants of panel A and B samples (3 h, 7 h and 24 h). The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. The right graph in panel D shows the comparison of the Western blot band intensity ratio between 3 h samples using the Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect any PFO production differences among these strains in 7 or 24 h samples (Figure S10B).

Techniques Used: Comparison, Mutagenesis, Cell Culture, Incubation, Western Blot, Software

Image Search Results

Journal: Toxins

Article Title: BrnQ Branched-Chain Amino Acid Transporters Influence Toxin Production by, but Not Growth of, Clostridium perfringens Type A Strain ATCC3624.

doi: 10.3390/toxins17040187

Figure Lengend Snippet: Figure 1. Expression of brnQ genes during culture in TY cultures of ATCC3624. (A) RT-PCR analyses. The recA gene was used as a housekeeping gene, and a sample lacking reverse transcriptase (RT-) was run (top panel) in a PCR reaction with recA primers to show the absence of DNA contamination in these RNA samples; recA gene RT-PCR (2nd panel) showed that cDNA was present in all samples. Further RT-PCR analyses (bottom three panels) are shown using brnQ1 primers (for brnQ1 expres- sion), brnQ2 primers (for brnQ2 expression), or brnQ3 primers (for brnQ3 expression) for wild-type ATCC3624 cultures at different time points. The expected size of each PCR product is listed in Table 1. The sample with ATCC3624 DNA served as the positive control, and the sample without ATCC3624 DNA served as the negative control. DNA size markers are shown on the left of each gel. The results shown are representative of three repetitions. (B) A comparison of the RT-PCR band intensity ratio between the brnQ gene vs. the housekeeping recA gene is shown using the Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the 2 h result.

Article Snippet: The current study used C. perfringens type A strain ATCC3624 (ATCC®, Manassas VA, USA) and E. coli DH5α competent cells [New England Biolabs (NEB), Ipswich, MA, USA].

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Reverse Transcription, Positive Control, Negative Control, Comparison, Software

Journal: Toxins

Article Title: BrnQ Branched-Chain Amino Acid Transporters Influence Toxin Production by, but Not Growth of, Clostridium perfringens Type A Strain ATCC3624.

doi: 10.3390/toxins17040187

Figure Lengend Snippet: Figure 2. A comparison of post-inoculation changes in culture OD600, vegetative cell viability, and PLC or PFO production levels for ATCC3624 (WT), its brnQ1 null mutant (brnQ1KO), and the complementing strain (brnQ1c) when cultured in a TY medium. (A) Post-inoculation changes in the OD600 of TY cultures of these strains from 0 to 9 h at 37 ◦C. The result shown is representative of three repetitions. (B) OD600 of TY cultures at 24 h at 37 ◦C (top). The same samples were also used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. (C) Western blot analyses of PLC level in supernatants of 3, 7, and 24 h culture samples (top panel). The PLC blot was also repeated using 10 µL more of the sample from the same 3 h TY sample, with longer film exposure (bottom left blot), to allow for a better comparison of PLC production between the brnQ1 null mutant vs. wild-type ATCC3624 or the complementing strain. The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio is shown between 3 h samples (bottom right graph) using Image J software. The graph shows the mean values from three independent experiments. The error bars indicate the S.D. * p < 0.05 relative to wild type. Similar Image J analyses did not detect PLC production differences among these strains in 7 or 24 h samples (Figure S5A). (D) Western blot analyses of the PFO level in supernatants of 3, 7, and 24 h culture samples. The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. Image J analyses did not detect significant differences in PFO production among these strains at any time points (Figure S5A).

Article Snippet: The current study used C. perfringens type A strain ATCC3624 (ATCC®, Manassas VA, USA) and E. coli DH5α competent cells [New England Biolabs (NEB), Ipswich, MA, USA].

Techniques: Comparison, Mutagenesis, Cell Culture, Western Blot, Software

Journal: Toxins

Article Title: BrnQ Branched-Chain Amino Acid Transporters Influence Toxin Production by, but Not Growth of, Clostridium perfringens Type A Strain ATCC3624.

doi: 10.3390/toxins17040187

Figure Lengend Snippet: Figure 3. A comparison of post-inoculation changes in culture OD600, vegetative cell viability, and PLC or PFO production levels for ATCC3624 (WT), its brnQ2 null mutant (brnQ2KO), and the complementing strain (brnQ2c) when cultured in TY medium. (A) Post-inoculation changes in the OD600 of TY cultures of these strains from 0 to 9 h at 37 ◦C. The result shown is representative of three repetitions. (B) The OD600 of TY cultures at 24 h at 37 ◦C (top). The same samples were used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. (C) Western blot analyses of PLC levels in supernatants of 3, 7, and 24 h culture samples (top panel). The PLC blot was also repeated using 10 µL more of the same 3 h TY samples, with longer film exposure (bottom left blot), to allow for a better comparison of PLC production between the brnQ2 null mutant vs. wild-type ATCC3624 or the complementing strain. The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio is shown between 3 h samples (bottom right graph) using Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect PLC production differences among the strains in 7 or 24 h samples (Figure S5B). (D) Western blot analyses of PFO levels in supernatants of 3, 7, and 24 h culture samples. The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown at left. Image J software did not detect significant differences in PFO production among the strains at any time points (Figure S5B).

Article Snippet: The current study used C. perfringens type A strain ATCC3624 (ATCC®, Manassas VA, USA) and E. coli DH5α competent cells [New England Biolabs (NEB), Ipswich, MA, USA].

Techniques: Comparison, Mutagenesis, Cell Culture, Western Blot, Software

Journal: Toxins

Article Title: BrnQ Branched-Chain Amino Acid Transporters Influence Toxin Production by, but Not Growth of, Clostridium perfringens Type A Strain ATCC3624.

doi: 10.3390/toxins17040187

Figure Lengend Snippet: Figure 4. A comparison of the post-inoculation changes in culture OD600, vegetative cell viability, and PLC or PFO production levels for ATCC3624 (WT), its brnQ3 null mutant (brnQ3KO), and the complementing strain (brnQ3c) when cultured in a TY medium. (A) Post-inoculation changes in the OD600 of TY cultures from 0 to 9 h at 37 ◦C. The result shown is representative of three repetitions. (B) The OD600 of TY cultures at 24 h at 37 ◦C (top). The same samples were also used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. (C) Western blot analyses of PLC levels in supernatants of 3, 7, and 24 h culture samples (top panel). The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio is shown between 3 h samples (bottom graph) using Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect PLC production differences among these strains in 7 or 24 h samples (Figure S5C). (D) Western blot analyses of PFO levels in supernatants of 3, 7, and 24 h culture samples. The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. Image J analyses did not detect significant differences in PFO production among these strains at any time points (Figure S5C).

Article Snippet: The current study used C. perfringens type A strain ATCC3624 (ATCC®, Manassas VA, USA) and E. coli DH5α competent cells [New England Biolabs (NEB), Ipswich, MA, USA].

Techniques: Comparison, Mutagenesis, Cell Culture, Western Blot, Software

Journal: Toxins

Article Title: BrnQ Branched-Chain Amino Acid Transporters Influence Toxin Production by, but Not Growth of, Clostridium perfringens Type A Strain ATCC3624.

doi: 10.3390/toxins17040187

Figure Lengend Snippet: Figure 5. A comparison of expression of the brnQ genes by wild-type ATCC3624, its isogenic brnQ null mutants, and complementing strains. ATCC3624, the single brnQ null mutants, or complementing strains were inoculated into TY broth and then incubated anaerobically for 2 h at 37 ◦C. Bacteria were collected and pelleted via centrifugation. Total RNA was extracted from the pellets, and cDNA was made for RT-qPCR analyses. Average CT values were normalized to the housekeeping recA gene, and the fold differences in expression were calculated using the comparative CT method (2−∆∆CT). The mean values from three independent experiments are shown. The error bars indicate the S.D. (A) RT- qPCR for the expression of brnQ2 or brnQ3 was performed using cDNA from the wild-type parent, the brnQ1 null mutant, and the complementing strain after their incubation in a TY medium for 2 h. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. (B) RT-qPCR for the expression of brnQ1 or brnQ3 was performed using cDNA from the wild type-parent, the brnQ2 null mutant, and the complementing strain after their incubation in TY medium for 2 h. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. (C) RT-qPCR for the expression of brnQ1 or brnQ2 was performed using cDNA from the wild-type parent, the brnQ3 null mutant, and the complementing strain after their incubation in TY medium for 2 h. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type.

Article Snippet: The current study used C. perfringens type A strain ATCC3624 (ATCC®, Manassas VA, USA) and E. coli DH5α competent cells [New England Biolabs (NEB), Ipswich, MA, USA].

Techniques: Comparison, Expressing, Incubation, Bacteria, Centrifugation, Quantitative RT-PCR, Mutagenesis

Journal: Toxins

Article Title: BrnQ Branched-Chain Amino Acid Transporters Influence Toxin Production by, but Not Growth of, Clostridium perfringens Type A Strain ATCC3624.

doi: 10.3390/toxins17040187

Figure Lengend Snippet: Figure 7. A comparison of PLC or PFO production levels for wild-type ATCC3624, its double null mutants, and the complementing strain when cultured in a TY medium. Western blot analyses of PLC (A) and PFO (B) levels in supernatants of the wild type, double null mutants, and their complementing strains in the early-stage culture (3 h). The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. The right figure in panels A and B show the comparison of Western blot intensities determined by Image J. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type.

Article Snippet: The current study used C. perfringens type A strain ATCC3624 (ATCC®, Manassas VA, USA) and E. coli DH5α competent cells [New England Biolabs (NEB), Ipswich, MA, USA].

Techniques: Comparison, Cell Culture, Western Blot

Journal: Toxins

Article Title: BrnQ Branched-Chain Amino Acid Transporters Influence Toxin Production by, but Not Growth of, Clostridium perfringens Type A Strain ATCC3624.

doi: 10.3390/toxins17040187

Figure Lengend Snippet: Figure 8. A comparison of post-inoculation changes in culture OD600, vegetative cell viability, or PLC and PFO production levels for ATCC3624 (WT) vs. the triple null mutant (TKO) when cultured in a TY medium. (A) Post-inoculation changes in the OD600 of TY cultures incubated for up to 9 h at 37 ◦C. The results shown are representative of three repetitions. (B) The OD600 of TY cultures after 24 h of incubation at 37 ◦C (top). The same samples were used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. (C) Western blot analyses of PLC levels in the supernatants of panel A and B samples (3 h, 7 h and 24 h). The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio between 3 h samples (right graph) using Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect any PLC production differences among these strains in 7 or 24 h samples (Figure S10A). (D) Western blot analyses of PFO levels in the supernatants of panel A and B samples (3 h, 7 h and 24 h). The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. The right graph in panel D shows the comparison of the Western blot band intensity ratio between 3 h samples using the Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect any PFO production differences among these strains in 7 or 24 h samples (Figure S10B).

Article Snippet: The current study used C. perfringens type A strain ATCC3624 (ATCC®, Manassas VA, USA) and E. coli DH5α competent cells [New England Biolabs (NEB), Ipswich, MA, USA].

Techniques: Comparison, Mutagenesis, Cell Culture, Incubation, Western Blot, Software

Expression of brnQ genes during culture in TY cultures of ATCC3624. ( A ) RT-PCR analyses. The recA gene was used as a housekeeping gene, and a sample lacking reverse transcriptase (RT-) was run (top panel) in a PCR reaction with recA primers to show the absence of DNA contamination in these RNA samples; recA gene RT-PCR (2nd panel) showed that cDNA was present in all samples. Further RT-PCR analyses (bottom three panels) are shown using brnQ1 primers (for brnQ1 expression), brnQ2 primers (for brnQ2 expression), or brnQ3 primers (for brnQ3 expression) for wild-type ATCC3624 cultures at different time points. The expected size of each PCR product is listed in . The sample with ATCC3624 DNA served as the positive control, and the sample without ATCC3624 DNA served as the negative control. DNA size markers are shown on the left of each gel. The results shown are representative of three repetitions. ( B ) A comparison of the RT-PCR band intensity ratio between the brnQ gene vs. the housekeeping recA gene is shown using the Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the 2 h result.

Journal: Toxins

Article Title: BrnQ Branched-Chain Amino Acid Transporters Influence Toxin Production by, but Not Growth of, Clostridium perfringens Type A Strain ATCC3624

doi: 10.3390/toxins17040187

Figure Lengend Snippet: Expression of brnQ genes during culture in TY cultures of ATCC3624. ( A ) RT-PCR analyses. The recA gene was used as a housekeeping gene, and a sample lacking reverse transcriptase (RT-) was run (top panel) in a PCR reaction with recA primers to show the absence of DNA contamination in these RNA samples; recA gene RT-PCR (2nd panel) showed that cDNA was present in all samples. Further RT-PCR analyses (bottom three panels) are shown using brnQ1 primers (for brnQ1 expression), brnQ2 primers (for brnQ2 expression), or brnQ3 primers (for brnQ3 expression) for wild-type ATCC3624 cultures at different time points. The expected size of each PCR product is listed in . The sample with ATCC3624 DNA served as the positive control, and the sample without ATCC3624 DNA served as the negative control. DNA size markers are shown on the left of each gel. The results shown are representative of three repetitions. ( B ) A comparison of the RT-PCR band intensity ratio between the brnQ gene vs. the housekeeping recA gene is shown using the Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the 2 h result.

Article Snippet: ATCC3624 , Wild type , Purchased from ATCC.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Reverse Transcription, Positive Control, Negative Control, Comparison, Software

A comparison of post-inoculation changes in culture OD 600 , vegetative cell viability, and PLC or PFO production levels for ATCC3624 (WT), its brnQ1 null mutant (brnQ1KO), and the complementing strain (brnQ1c) when cultured in a TY medium. ( A ) Post-inoculation changes in the OD 600 of TY cultures of these strains from 0 to 9 h at 37 °C. The result shown is representative of three repetitions. ( B ) OD 600 of TY cultures at 24 h at 37 °C (top). The same samples were also used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. ( C ) Western blot analyses of PLC level in supernatants of 3, 7, and 24 h culture samples (top panel). The PLC blot was also repeated using 10 µL more of the sample from the same 3 h TY sample, with longer film exposure (bottom left blot), to allow for a better comparison of PLC production between the brnQ1 null mutant vs. wild-type ATCC3624 or the complementing strain. The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio is shown between 3 h samples (bottom right graph) using Image J software. The graph shows the mean values from three independent experiments. The error bars indicate the S.D. * p < 0.05 relative to wild type. Similar Image J analyses did not detect PLC production differences among these strains in 7 or 24 h samples . ( D ) Western blot analyses of the PFO level in supernatants of 3, 7, and 24 h culture samples. The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. Image J analyses did not detect significant differences in PFO production among these strains at any time points .

Journal: Toxins

Article Title: BrnQ Branched-Chain Amino Acid Transporters Influence Toxin Production by, but Not Growth of, Clostridium perfringens Type A Strain ATCC3624

doi: 10.3390/toxins17040187

Figure Lengend Snippet: A comparison of post-inoculation changes in culture OD 600 , vegetative cell viability, and PLC or PFO production levels for ATCC3624 (WT), its brnQ1 null mutant (brnQ1KO), and the complementing strain (brnQ1c) when cultured in a TY medium. ( A ) Post-inoculation changes in the OD 600 of TY cultures of these strains from 0 to 9 h at 37 °C. The result shown is representative of three repetitions. ( B ) OD 600 of TY cultures at 24 h at 37 °C (top). The same samples were also used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. ( C ) Western blot analyses of PLC level in supernatants of 3, 7, and 24 h culture samples (top panel). The PLC blot was also repeated using 10 µL more of the sample from the same 3 h TY sample, with longer film exposure (bottom left blot), to allow for a better comparison of PLC production between the brnQ1 null mutant vs. wild-type ATCC3624 or the complementing strain. The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio is shown between 3 h samples (bottom right graph) using Image J software. The graph shows the mean values from three independent experiments. The error bars indicate the S.D. * p < 0.05 relative to wild type. Similar Image J analyses did not detect PLC production differences among these strains in 7 or 24 h samples . ( D ) Western blot analyses of the PFO level in supernatants of 3, 7, and 24 h culture samples. The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. Image J analyses did not detect significant differences in PFO production among these strains at any time points .

Article Snippet: ATCC3624 , Wild type , Purchased from ATCC.

Techniques: Comparison, Mutagenesis, Cell Culture, Western Blot, Software

A comparison of post-inoculation changes in culture OD 600 , vegetative cell viability, and PLC or PFO production levels for ATCC3624 (WT), its brnQ2 null mutant (brnQ2KO), and the complementing strain (brnQ2c) when cultured in TY medium. ( A ) Post-inoculation changes in the OD 600 of TY cultures of these strains from 0 to 9 h at 37 °C. The result shown is representative of three repetitions. ( B ) The OD 600 of TY cultures at 24 h at 37 °C (top). The same samples were used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. ( C ) Western blot analyses of PLC levels in supernatants of 3, 7, and 24 h culture samples (top panel). The PLC blot was also repeated using 10 µL more of the same 3 h TY samples, with longer film exposure (bottom left blot), to allow for a better comparison of PLC production between the brnQ2 null mutant vs. wild-type ATCC3624 or the complementing strain. The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio is shown between 3 h samples (bottom right graph) using Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect PLC production differences among the strains in 7 or 24 h samples . ( D ) Western blot analyses of PFO levels in supernatants of 3, 7, and 24 h culture samples. The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown at left. Image J software did not detect significant differences in PFO production among the strains at any time points .

Journal: Toxins

Article Title: BrnQ Branched-Chain Amino Acid Transporters Influence Toxin Production by, but Not Growth of, Clostridium perfringens Type A Strain ATCC3624

doi: 10.3390/toxins17040187

Figure Lengend Snippet: A comparison of post-inoculation changes in culture OD 600 , vegetative cell viability, and PLC or PFO production levels for ATCC3624 (WT), its brnQ2 null mutant (brnQ2KO), and the complementing strain (brnQ2c) when cultured in TY medium. ( A ) Post-inoculation changes in the OD 600 of TY cultures of these strains from 0 to 9 h at 37 °C. The result shown is representative of three repetitions. ( B ) The OD 600 of TY cultures at 24 h at 37 °C (top). The same samples were used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. ( C ) Western blot analyses of PLC levels in supernatants of 3, 7, and 24 h culture samples (top panel). The PLC blot was also repeated using 10 µL more of the same 3 h TY samples, with longer film exposure (bottom left blot), to allow for a better comparison of PLC production between the brnQ2 null mutant vs. wild-type ATCC3624 or the complementing strain. The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio is shown between 3 h samples (bottom right graph) using Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect PLC production differences among the strains in 7 or 24 h samples . ( D ) Western blot analyses of PFO levels in supernatants of 3, 7, and 24 h culture samples. The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown at left. Image J software did not detect significant differences in PFO production among the strains at any time points .

Article Snippet: ATCC3624 , Wild type , Purchased from ATCC.

Techniques: Comparison, Mutagenesis, Cell Culture, Western Blot, Software

A comparison of the post-inoculation changes in culture OD 600 , vegetative cell viability, and PLC or PFO production levels for ATCC3624 (WT), its brnQ3 null mutant (brnQ3KO), and the complementing strain (brnQ3c) when cultured in a TY medium. ( A ) Post-inoculation changes in the OD 600 of TY cultures from 0 to 9 h at 37 °C. The result shown is representative of three repetitions. ( B ) The OD 600 of TY cultures at 24 h at 37 °C (top). The same samples were also used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. ( C ) Western blot analyses of PLC levels in supernatants of 3, 7, and 24 h culture samples (top panel). The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio is shown between 3 h samples (bottom graph) using Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect PLC production differences among these strains in 7 or 24 h samples . ( D ) Western blot analyses of PFO levels in supernatants of 3, 7, and 24 h culture samples. The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. Image J analyses did not detect significant differences in PFO production among these strains at any time points .

Journal: Toxins

Article Title: BrnQ Branched-Chain Amino Acid Transporters Influence Toxin Production by, but Not Growth of, Clostridium perfringens Type A Strain ATCC3624

doi: 10.3390/toxins17040187

Figure Lengend Snippet: A comparison of the post-inoculation changes in culture OD 600 , vegetative cell viability, and PLC or PFO production levels for ATCC3624 (WT), its brnQ3 null mutant (brnQ3KO), and the complementing strain (brnQ3c) when cultured in a TY medium. ( A ) Post-inoculation changes in the OD 600 of TY cultures from 0 to 9 h at 37 °C. The result shown is representative of three repetitions. ( B ) The OD 600 of TY cultures at 24 h at 37 °C (top). The same samples were also used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. ( C ) Western blot analyses of PLC levels in supernatants of 3, 7, and 24 h culture samples (top panel). The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio is shown between 3 h samples (bottom graph) using Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect PLC production differences among these strains in 7 or 24 h samples . ( D ) Western blot analyses of PFO levels in supernatants of 3, 7, and 24 h culture samples. The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. Image J analyses did not detect significant differences in PFO production among these strains at any time points .

Article Snippet: ATCC3624 , Wild type , Purchased from ATCC.

Techniques: Comparison, Mutagenesis, Cell Culture, Western Blot, Software

A comparison of expression of the brnQ genes by wild-type ATCC3624, its isogenic brnQ null mutants, and complementing strains. ATCC3624, the single brnQ null mutants, or complementing strains were inoculated into TY broth and then incubated anaerobically for 2 h at 37 °C. Bacteria were collected and pelleted via centrifugation. Total RNA was extracted from the pellets, and cDNA was made for RT-qPCR analyses. Average C T values were normalized to the housekeeping recA gene, and the fold differences in expression were calculated using the comparative C T method (2 −ΔΔC T ). The mean values from three independent experiments are shown. The error bars indicate the S.D. ( A ) RT-qPCR for the expression of brnQ2 or brnQ3 was performed using cDNA from the wild-type parent, the brnQ1 null mutant, and the complementing strain after their incubation in a TY medium for 2 h. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. ( B ) RT-qPCR for the expression of brnQ1 or brnQ3 was performed using cDNA from the wild type-parent, the brnQ2 null mutant, and the complementing strain after their incubation in TY medium for 2 h. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. ( C ) RT-qPCR for the expression of brnQ1 or brnQ2 was performed using cDNA from the wild-type parent, the brnQ3 null mutant, and the complementing strain after their incubation in TY medium for 2 h. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type.

Journal: Toxins

Article Title: BrnQ Branched-Chain Amino Acid Transporters Influence Toxin Production by, but Not Growth of, Clostridium perfringens Type A Strain ATCC3624

doi: 10.3390/toxins17040187

Figure Lengend Snippet: A comparison of expression of the brnQ genes by wild-type ATCC3624, its isogenic brnQ null mutants, and complementing strains. ATCC3624, the single brnQ null mutants, or complementing strains were inoculated into TY broth and then incubated anaerobically for 2 h at 37 °C. Bacteria were collected and pelleted via centrifugation. Total RNA was extracted from the pellets, and cDNA was made for RT-qPCR analyses. Average C T values were normalized to the housekeeping recA gene, and the fold differences in expression were calculated using the comparative C T method (2 −ΔΔC T ). The mean values from three independent experiments are shown. The error bars indicate the S.D. ( A ) RT-qPCR for the expression of brnQ2 or brnQ3 was performed using cDNA from the wild-type parent, the brnQ1 null mutant, and the complementing strain after their incubation in a TY medium for 2 h. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. ( B ) RT-qPCR for the expression of brnQ1 or brnQ3 was performed using cDNA from the wild type-parent, the brnQ2 null mutant, and the complementing strain after their incubation in TY medium for 2 h. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. ( C ) RT-qPCR for the expression of brnQ1 or brnQ2 was performed using cDNA from the wild-type parent, the brnQ3 null mutant, and the complementing strain after their incubation in TY medium for 2 h. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type.

Article Snippet: ATCC3624 , Wild type , Purchased from ATCC.

Techniques: Comparison, Expressing, Incubation, Bacteria, Centrifugation, Quantitative RT-PCR, Mutagenesis

A comparison of PLC or PFO production levels for wild-type ATCC3624, its double null mutants, and the complementing strain when cultured in a TY medium. Western blot analyses of PLC ( A ) and PFO ( B ) levels in supernatants of the wild type, double null mutants, and their complementing strains in the early-stage culture (3 h). The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. The right figure in panels A and B show the comparison of Western blot intensities determined by Image J. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type.

Journal: Toxins

Article Title: BrnQ Branched-Chain Amino Acid Transporters Influence Toxin Production by, but Not Growth of, Clostridium perfringens Type A Strain ATCC3624

doi: 10.3390/toxins17040187

Figure Lengend Snippet: A comparison of PLC or PFO production levels for wild-type ATCC3624, its double null mutants, and the complementing strain when cultured in a TY medium. Western blot analyses of PLC ( A ) and PFO ( B ) levels in supernatants of the wild type, double null mutants, and their complementing strains in the early-stage culture (3 h). The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. The right figure in panels A and B show the comparison of Western blot intensities determined by Image J. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type.

Article Snippet: ATCC3624 , Wild type , Purchased from ATCC.

Techniques: Comparison, Cell Culture, Western Blot

A comparison of post-inoculation changes in culture OD 600 , vegetative cell viability, or PLC and PFO production levels for ATCC3624 (WT) vs. the triple null mutant (TKO) when cultured in a TY medium. ( A ) Post-inoculation changes in the OD 600 of TY cultures incubated for up to 9 h at 37 °C. The results shown are representative of three repetitions. ( B ) The OD 600 of TY cultures after 24 h of incubation at 37 °C (top). The same samples were used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. ( C ) Western blot analyses of PLC levels in the supernatants of panel A and B samples (3 h, 7 h and 24 h). The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio between 3 h samples (right graph) using Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect any PLC production differences among these strains in 7 or 24 h samples . ( D ) Western blot analyses of PFO levels in the supernatants of panel A and B samples (3 h, 7 h and 24 h). The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. The right graph in panel D shows the comparison of the Western blot band intensity ratio between 3 h samples using the Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect any PFO production differences among these strains in 7 or 24 h samples .

Journal: Toxins

Article Title: BrnQ Branched-Chain Amino Acid Transporters Influence Toxin Production by, but Not Growth of, Clostridium perfringens Type A Strain ATCC3624

doi: 10.3390/toxins17040187

Figure Lengend Snippet: A comparison of post-inoculation changes in culture OD 600 , vegetative cell viability, or PLC and PFO production levels for ATCC3624 (WT) vs. the triple null mutant (TKO) when cultured in a TY medium. ( A ) Post-inoculation changes in the OD 600 of TY cultures incubated for up to 9 h at 37 °C. The results shown are representative of three repetitions. ( B ) The OD 600 of TY cultures after 24 h of incubation at 37 °C (top). The same samples were used for colony counting of viable vegetative cells (bottom). The mean values from three independent experiments are shown. The error bars indicate the S.D. ( C ) Western blot analyses of PLC levels in the supernatants of panel A and B samples (3 h, 7 h and 24 h). The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. A comparison of the Western blot band intensity ratio between 3 h samples (right graph) using Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect any PLC production differences among these strains in 7 or 24 h samples . ( D ) Western blot analyses of PFO levels in the supernatants of panel A and B samples (3 h, 7 h and 24 h). The results shown are representative of three repetitions. The size of proteins in kiloDaltons (kDa) is shown on the left. The right graph in panel D shows the comparison of the Western blot band intensity ratio between 3 h samples using the Image J software. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. Similar analyses did not detect any PFO production differences among these strains in 7 or 24 h samples .

Article Snippet: ATCC3624 , Wild type , Purchased from ATCC.

Techniques: Comparison, Mutagenesis, Cell Culture, Incubation, Western Blot, Software

C. perfringens strains used in this study.

Journal: Toxins

Article Title: BrnQ Branched-Chain Amino Acid Transporters Influence Toxin Production by, but Not Growth of, Clostridium perfringens Type A Strain ATCC3624

doi: 10.3390/toxins17040187

Figure Lengend Snippet: C. perfringens strains used in this study.

Article Snippet: ATCC3624 , Wild type , Purchased from ATCC.

Techniques: Mutagenesis

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