Investigating a Viral Response Gene Using the CRISPR Editing Process
Faculty Sponsor(s)
Jennifer Grier
Location
Jereld R. Nicholson Library: Grand Avenue
Subject Area
Biochemistry
Description
Of many genes that respond to viral infections, most are not well understood. HP, a gene normally involved in hemoglobin recycling, is known to be upregulated after viral infection. In an effort to mutate this gene of interest and better understand the function of this gene, the CRISPR-Cas9 system was used. CRISPR is a gene editing tool that cleaves the targeted DNA of the gene, potentially causing a mutation. A mutation or knockout of the gene could illustrate how the immune response functions in the absence of this upregulated gene. This process was done with two guides (sgRNA) to increase the chances of knocking out the gene and because we had reason to believe both were equally useful. We are certain that the plasmids contain our guides due to results from gel electrophoresis and confirmation by PCR sequencing; we also found that the cells express the plasmid because the cells survived antibiotic selection processes. Primers were designed for both PCR genomic sequencing and confirmation of the mutation. In the future, sequencing and analysis of the DNA and proteins can be done to confirm mutation of HP.
Recommended Citation
Vanderschaegen, Anna; Reece, Shae; and Rempelos, Sarah, "Investigating a Viral Response Gene Using the CRISPR Editing Process" (2017). Linfield University Student Symposium: A Celebration of Scholarship and Creative Achievement. Event. Submission 49.
https://digitalcommons.linfield.edu/symposium/2017/all/49
Investigating a Viral Response Gene Using the CRISPR Editing Process
Jereld R. Nicholson Library: Grand Avenue
Of many genes that respond to viral infections, most are not well understood. HP, a gene normally involved in hemoglobin recycling, is known to be upregulated after viral infection. In an effort to mutate this gene of interest and better understand the function of this gene, the CRISPR-Cas9 system was used. CRISPR is a gene editing tool that cleaves the targeted DNA of the gene, potentially causing a mutation. A mutation or knockout of the gene could illustrate how the immune response functions in the absence of this upregulated gene. This process was done with two guides (sgRNA) to increase the chances of knocking out the gene and because we had reason to believe both were equally useful. We are certain that the plasmids contain our guides due to results from gel electrophoresis and confirmation by PCR sequencing; we also found that the cells express the plasmid because the cells survived antibiotic selection processes. Primers were designed for both PCR genomic sequencing and confirmation of the mutation. In the future, sequencing and analysis of the DNA and proteins can be done to confirm mutation of HP.
