Pseudomonas frederiksbergensis is a gram-negative bacteria. It has been found by Andersen and team (2000) in Frederiksberg, Copenhagen, Denmark at a coal gasification site.
- Andersen S, Johnsen K, Sorensen J, Nielsen P, Jacobsen C. 2000. Pseudomonas frederiksbergensis sp. nov., isolated from soil at a coal gasification site. Int. J. of Syst. and Evol. Microbiol. 50:1957-1964.
Date collected: February 9, 2017
Methods for isolation and identification:
- Dry soil 4 meters away from Buffalo Creek in Lancer Park (Figure 1). Half of a gram of the soil was added to 50 milliliters of sterile water. After vortexing and resting, the bacteria were diluted using a serial dilution then plated onto an LB agar plate. The plate was then incubated at room temperature for 18-24 hours.
- A pale yellow circular colony (Figure 2) was selected from the 1:100 count plate. This colony was used for the PCR amplification.
- The genus and species of the bacteria was identified using PCR product that was digested with MspI.
Figure 1. Site of collection
Figure 2. Selected colony
- MspI digestion (Figure 3): A 2,000 bp product was amplified by PCR. Three bands were identified at approximately 700 bp, 450 bp, and 150 bp after MspI digestion and gel electrophoresis.
Figure 3. Results of MspI digestion
- Sequence analysis (Figures 4 & 5): The sequenced PCR product generated 614 bases of high-quality reads that were used to identify the genus and species of the colony. The chromatogram of the sequence is available in Figure 4. The NCBI BLAST analysis 99% identity in base pairs 74-688 of the 16s rRNA gene of Pseudomonas frederiksbergensis.
Figure 4. Chromatogram of Pseudomonas frederiksbergensis
Figure 5. NCBI Blast Analysis
Contributed by: Kathryn Kosiorek and Claire Walker, BIOL 250 Spring 2017, Group 15