Researcher(s)
- Matthew Douglass, Biological Sciences, University of Delaware
Faculty Mentor(s)
- Jeremy Bird, Department of Biological Sciences, University of Delaware
Abstract
A growing problem in the field of medicine is that of antibiotic resistant bacteria. A potential solution to this issue is the use of bacteriophages, viruses that infect bacteria, to treat bacterial infections. Through evolution, bacteria have developed multiple ways of protecting themselves against bacteriophage infection. The Clustered Regularly Interspaced Short Palindromic Repeats(CRISPR)-Cas system is one of these, and if present in pathogenic bacteria, this system could cause issues for bacteriophage therapy techniques. The CRISPR system allows the cell to recognize foreign DNA and RNA sequences that it has encountered before and destroy them.
In this study, I performed CRISPR protection assays to test how well a Type III-A CRISPR system, taken from the bacteria S. epidermidis, can protect E. coli from infection by the T4 and T4R bacteriophages. The Type III-A CRISPR system targets bacteriophage RNA while it is being transcribed and destroys both the target RNA and the DNA that it is being transcribed from. This system, expressable off of a plasmid, can be altered to target any gene. The plasmids used in this study were generated by students in BISC430: Experimental Microbiology, containing targeting spacers against genes from the T4 bacteriophage. I tested all 12 E. coli plasmids containing targeting spacers for genes of interest from the T4 bacteriophage and the genes encoding the CRISPR-Cas Type III-A system to protect E. coli from bacteriophage infection. I found that targeting certain genes, specifically denB.1, denV, and A-gt, protects against infection more than the rest, possibly due to their vital roles in the process of bacteriophage infection. By learning more about bacteriophages, we may be able to weaponize them to fight off bacterial infections in the future.