Isolation and Characterization of Novel Mycobacteriophages From the Central Illinois Region

Members of the Illinois Wesleyan General Biology Science Education Alliance (SEA) laboratory isolated and characterized fifteen distinctive phages capable of infecting Mycobacterium smegmatis. Each student collected soil samples from the central Illinois area and used direct plating or enrichment techniques to isolate phages. Streak assays were used to purify single phage populations. Individual phage populations were then characterized and DNA was isolated. Based on the following characteristics; plaque morphology (size and turbidity), life style (temperate or lytic) and DNA restriction patterns, we determined that each student has isolated a unique phage. The DNA from a single Mycobacteriophage, Kazan, was sent to the University of Pittsburgh for genome sequencing. DNA Sequencing determined that Kazan is 52,160 base pairs, including 10 base pair 3\u27 overhang (CGGTCGGTTA), and a member of the A6 subcluster of Mycobacteriophages. Kazan is most closely related to the phages EricB and DaVinci (99% identity). Genome analysis, using the computer programs DNA Master, Glimmer, GeneMark, and Aragorn, determined that the Kazan genome housed 99 genes and 3 tRNAs. The potential protein function for each gene was determined using the computer programs HHPred, BLASTP and Phamerator. All the individual phage data was submitted to the Mycobacteriophage DataBase and the genome annotation, when completed, will be submitted to the DNA database, GenBank

The Isolation and Characterization of Novel Bacteriophages from Central Illinois

Members of the Illinois Wesleyan University General Biology Science Education Alliance (SEA) laboratory isolated and characterized a number of distinctive mycobacteriophages. Each student collected soil samples from the central Illinois area then used streak assays and titrations to isolate unique phages that infect Mycobacterium smegmatis. Transmission electron microscopy (TEM) and DNA restriction enzyme digests were used to determine the morphology and tentative cluster placement of each phage. By the end of the semester, fifteen novel phages were isolated with a wide range of characteristics, despite the small sampling area. This data was submitted to the Mycobacteriophage DataBase. After analysis and class discussion, the DNA from three different phages were sent to the University of Pittsburgh for genome sequencing