Temperate Phage Isolated from Potted Plant Soil Grace

Temperate Phage Isolated from Potted Plant Soil Grace Perkins and Katherine Jacobsen Phage Genomics Lab Section 014 University of Colorado, Boulder March 15, 2020

Phage Life Cycles and Other Information

Temperate Phage Identified After Streaking Large, cloudy plaques with an average diameter of 5 cm Plaques purified through seven rounds of titer assays with a final titer of 3. 7 X 10^10 pfu/m. L

Sal Hind III Hae III Eco RI Cla I Bam HI Uncut DNA 1 kb DNA Ladder Phage Isolated has Similar Restriction Digest to Phage Jasper in Cluster A, Subcluster A 1 10 KB 8 KB 6 KB 5 KB 4 KB 3 KB 2 KB 1. 5 KB 1. 0 KB 0. 5 KB Experimentally Isolated Phage, ~24 KB Jasper, A 1

A 3 - A 3+ N- N? A 1 - ● Restriction digest matches others A 1? ● Turbid plaques 1 kb DNA Ladder The Phage isolated is temperate with an indeterminate cluster from Cluster A, Subcluster A 1 ● PCR results inconclusive ○ No bands, even for controls ○ Error unknown ● Next steps: rerun PCR, electron microscopy, phage therapy or bioengineering PCR gel for isolated phage (A 3+ and A 3 - were total controls)

Phage Therapy: Going Temperate? ● Experiment: genetically modifying naturally temperate phage through allelic exchange with the goal of inducing a fully lytic life cycle. ● Results: Some phage were able to be engineered to increase the lytic activity ○ Specifically- ΦEf 11 (enterococcus faecalis), B 025 (listeria monocytogenes), and PSA (listeria monocytogenes)

References Phage Therapy: Going Temperate? https: //www. ncbi. nlm. nih. gov/pubmed/30466900 Barr Laboratory | Research Areas https: //thebarrlab. org/research. php? area=Bioengineering+Phages