Using Microbacterium foliorum as a Host for the Isolation and Subsequent Annotation of a Novel Bacteriophage Genome
College
College of Arts and Sciences
Department
Biology
Faculty Mentor
Victoria Frost, Ph.D., and Kristi Westover, Ph.D.
Abstract
Bacteriophages are viruses that inject their genomes into specific bacterial hosts to replicate. As members of the Howard Hughes Medical Institute’s SEA-PHAGES (Science Education Alliance – Phage Hunters Advancing Genomics and Evolutionary Science) program, freshman at Winthrop University have isolated 21 phages from soil using the host bacterial strain Mycobacteria smegmatis. This semester, the host bacterial strain Microbacterium foliorum was piloted to increase the collection and knowledge of diversity within actinobacteriophage in Rock Hill. The novel phage Scamander was isolated directly from local soil. The subsequent purification and amplification protocols entailed multiple plaque assays, which made use of techniques such as picking a plaque from plates and various dilutions of phage lysate to obtain a pure phage sample with a high titer. Transmission electron microscopy showed that Scamander has a long, flexible tail, which is characteristic of the siphoviridae morphotype. Phage DNA was extracted and the genome was sequenced by the Pittsburgh Bacteriophage Institute. Annotation of the genome is in progress and the following programs are being used to determine specific gene data: protein blasts from PhagesDB, HHPred, and NCBI; gene start-site calling programs such as Starterator, GeneMaster, and Glimmer; and genome comparison maps on Phamerator. When complete, the annotated genome will reveal functions of certain genes, while also serving as a comparison for other existing and, as yet, undiscovered Microbacterium phages. The increasing database of bacteriophages and their characteristics helps to expand the understanding of the genomic diversity of viruses, particularly those that infect bacterial hosts in the phylum Actinobacteria.
Course Assignment
BIOL 471 – Westover
Previously Presented/Performed?
Association of Southeastern Biologists (ASB) Annual Meeting, Myrtle Beach, South Carolina, March 2018
Start Date
20-4-2018 2:45 PM
Using Microbacterium foliorum as a Host for the Isolation and Subsequent Annotation of a Novel Bacteriophage Genome
West 221
Bacteriophages are viruses that inject their genomes into specific bacterial hosts to replicate. As members of the Howard Hughes Medical Institute’s SEA-PHAGES (Science Education Alliance – Phage Hunters Advancing Genomics and Evolutionary Science) program, freshman at Winthrop University have isolated 21 phages from soil using the host bacterial strain Mycobacteria smegmatis. This semester, the host bacterial strain Microbacterium foliorum was piloted to increase the collection and knowledge of diversity within actinobacteriophage in Rock Hill. The novel phage Scamander was isolated directly from local soil. The subsequent purification and amplification protocols entailed multiple plaque assays, which made use of techniques such as picking a plaque from plates and various dilutions of phage lysate to obtain a pure phage sample with a high titer. Transmission electron microscopy showed that Scamander has a long, flexible tail, which is characteristic of the siphoviridae morphotype. Phage DNA was extracted and the genome was sequenced by the Pittsburgh Bacteriophage Institute. Annotation of the genome is in progress and the following programs are being used to determine specific gene data: protein blasts from PhagesDB, HHPred, and NCBI; gene start-site calling programs such as Starterator, GeneMaster, and Glimmer; and genome comparison maps on Phamerator. When complete, the annotated genome will reveal functions of certain genes, while also serving as a comparison for other existing and, as yet, undiscovered Microbacterium phages. The increasing database of bacteriophages and their characteristics helps to expand the understanding of the genomic diversity of viruses, particularly those that infect bacterial hosts in the phylum Actinobacteria.
