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1.
FIG. 6.

FIG. 6. From: Regulated Expression of the Beta2-Toxin Gene (cpb2) in Clostridium perfringens Type A Isolates from Horses with Gastrointestinal Diseases.

Overproduction of CPB2 in horse GI disease isolates with a recombinant plasmid carrying cpb2 from 106902. Culture supernatant proteins, prepared from each of the specified C. perfringens isolates, were subjected to Western blot analysis. The blot was probed with CPB2 antibodies and developed by chemiluminescent detection to identify immunoreactive species. Molecular mass markers are shown on the left.

Michael Waters, et al. J Clin Microbiol. 2005 August;43(8):4002-4009.
2.
FIG. 2.

FIG. 2. From: Regulated Expression of the Beta2-Toxin Gene (cpb2) in Clostridium perfringens Type A Isolates from Horses with Gastrointestinal Diseases.

RFLP-Southern blot analysis of HpaI- or EcoRV-digested DNA from horse GI disease isolates. Total DNA isolated from each of the specified C. perfringens strains was digested with HpaI (A) or EcoRV (B) and then Southern transferred. The Southern blots were probed with a 318-bp DIG-labeled cpb2-specific probe. Results shown for control isolates include those for 106527 (a cpb2-negative type C strain). Representative cpb2-positive horse GI disease isolates include AHT2600, AHT2779, AHT2911, AHT3553, and AHT3992. Migration of the hybridizing band derived from each strain is indicated between the two blots.

Michael Waters, et al. J Clin Microbiol. 2005 August;43(8):4002-4009.
3.
FIG. 4.

FIG. 4. From: Regulated Expression of the Beta2-Toxin Gene (cpb2) in Clostridium perfringens Type A Isolates from Horses with Gastrointestinal Diseases.

RT-PCR analysis of C. perfringens strains. (A) Total RNA prepared from each specified strain was subjected to RT-PCR analysis using cpb2-specific internal primers as described in Materials and Methods. RT (+) and RT (−) indicate the presence and absence, respectively, of RT in the PCR. The RT-PCR-amplified products were analyzed by agarose (1%) gel electrophoresis and photographed under UV light. Molecular sizes of the DNA markers are given on the right. (B) Total DNA isolated from each specified strain was subjected to PCR analysis using the same cpb2-specific internal primers as those used for RT-PCR. The PCR-amplified products were analyzed by agarose (1%) gel electrophoresis and photographed under UV light. Molecular sizes of the DNA markers are given on the left.

Michael Waters, et al. J Clin Microbiol. 2005 August;43(8):4002-4009.
4.
FIG. 3.

FIG. 3. From: Regulated Expression of the Beta2-Toxin Gene (cpb2) in Clostridium perfringens Type A Isolates from Horses with Gastrointestinal Diseases.

PFGE evidence supporting the plasmid localization of cpb2 in horse GI disease isolates. PFGE and Southern hybridizing analysis of undigested DNA, prepared in agarose plugs, from each of the specified C. perfringens isolates. Blots were probed with a 318-bp cpb2-specific probe. Results shown for control isolates include those for CWC245 (a cpb2-positive type C isolate carrying the cpb2 gene on a large plasmid) and 106527 (a cpb2-negative type C isolate). Representative results shown for cpb2 horse GI disease isolates include those for 106902, 106903, AHT3992, and AHT4578. The pulsed-field gel was calibrated with λ DNA markers, whose migration is shown at the left of the blot.

Michael Waters, et al. J Clin Microbiol. 2005 August;43(8):4002-4009.
5.
FIG. 1.

FIG. 1. From: Regulated Expression of the Beta2-Toxin Gene (cpb2) in Clostridium perfringens Type A Isolates from Horses with Gastrointestinal Diseases.

Western blot analysis of CPB2 production by selected horse GI disease isolates. (A) Culture supernatant (sup) or total cell (cell) proteins, prepared from each of the specified C. perfringens isolates, were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and stained with Coomassie brilliant blue. Migration of the supernatant proteins is shown by arrows. (B) Western blot of the gel shown in panel A. The blot was probed with CPB2 antibodies and developed by chemiluminescent detection to identify immunoreactive species. Results shown for control isolates include those for CWC245 (a cpb2-positive type C strain). Results shown for representative horse GI disease isolates include those for isolates 106902 and 106903. Molecular mass markers are shown in the middle.

Michael Waters, et al. J Clin Microbiol. 2005 August;43(8):4002-4009.
6.
FIG. 5.

FIG. 5. From: Regulated Expression of the Beta2-Toxin Gene (cpb2) in Clostridium perfringens Type A Isolates from Horses with Gastrointestinal Diseases.

Northern blot analysis of cpb2 mRNAs from C. perfringens. (A) Expression of cpb2 mRNAs from horse GI disease isolates. RNA was extracted from specified C. perfringens cultures grown at 37°C for 3 h, and 8 μg of RNA from each was subjected to Northern blot analysis using a 560-bp alkaline phosphatase-labeled cpb2-specific probe. Results shown for control isolates include those for CWC245 (a known cpb2-positive type C strain) and 106527 (a cpb2-negative type C strain). Representative results shown for cpb2-positive horse GI disease isolates include those for 106902 and 106903. The cpb2 mRNA (1.2 kb) is indicated by an arrow. (B) Time course expression of cpb2 mRNAs. RNA was extracted from CWC245 and 106903 cultures grown at 37°C for specified time periods, and 8 μg of RNA from each was subjected to Northern blot analysis using a cpb2-specific probe. The cpb2 mRNA (1.2 kb) is indicated by arrowheads. (C) Comparative expression of cpb2 mRNAs from representative horse GI disease isolates and that from CWC245. Various amounts of RNA (0.1 to 8 μg), extracted from CWC245 (0.1 to 5 μg), 106902 (8 μg), and 106903 (8 μg) cultures grown at 37°C for 3 h, were subjected to Northern blot analysis using a 560-bp alkaline phosphatase-labeled cpb2-specific probe. The relative levels of cpb2 mRNA in horse GI disease isolates were determined as described in Materials and Methods. The cpb2 mRNA (1.2 kb) is indicated by an arrowhead. The various amounts of RNA loaded onto the gel are indicated at the bottom of the blot.

Michael Waters, et al. J Clin Microbiol. 2005 August;43(8):4002-4009.

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