Date of Graduation
8-2019
Document Type
Thesis
Degree Name
Bachelor of Science in Biomedical Engineering
Degree Level
Undergraduate
Department
Biomedical Engineering
Advisor/Mentor
Muldoon, Timothy
Abstract
CCL2 is an inflammatory mediator that is released by tumor cells to activate and direct immune cell species, especially macrophages, to inflammatory sites within the body. The goal of this project was to successfully generate knockout the CCL2 ligand gene using a CRISPR/Cas9 complex delivered via lipid mediated transfection. The sgRNA and Cas9 mRNA were introduced into the cells via lipid-mediated transfection. The cells were incubated for 4 days, before being analyzed using PCR and gel electrophoresis. We expected to see one band on the first gel and two bands on the second gel. Two bands appeared on the first gel and 2 bands appeared on the second gel. This indicates that the target sequence was amplified, but the transfection efficiency was too low for the enzyme to detect cleavage. We conclude that lipid mediated transfection is an efficient method for introducing foreign genetic material into CT-26 cells.
Keywords
CCL2, CRIPSR, Cas9, transfection, sgRNA
Citation
Sullivan, E. (2019). Generation of a CCL2 knockout using CRISPR/CAS9 and lipid mediated transfection in CT-26 murine colon carcinoma cells. Biomedical Engineering Undergraduate Honors Theses Retrieved from https://scholarworks.uark.edu/bmeguht/76