Submission Title

CRISPR-Cas9 Induced Mutations of IFI27 Gene

Location

Jereld R. Nicholson Library: Grand Avenue

Subject Area

Biochemistry

Description

IFI27 is a gene that has been previously associated with lung clear cell sarcoma as well as Newcastle disease. The importance of the IFI27 gene was explored using mutation inducing procedures involving the CRISPR-Cas9. CRISPR-Cas9 was discovered as defense mechanism in bacterial cells against viral infections. CRISPR-Cas9 is a gene editing technique that is used to cut out a gene of interest, and this is essential in determining how important the IFI27 gene is in cellular function. To target the IFI27 gene, a guide RNA sequence was chosen from important segments of the gene. This guide RNA sequence was inserted into a plasmid called pSpCas9 obtained by MIT. These modified plasmids were purified using antibacterial selection with LB/ampicillin plates. The purified plasmids were characterized using primers and PCR to amplify our desired guide RNA sequence and analyzed using gel electrophoresis to conclude our insertion was successful. Successful insertions promoted the decision to amplify our plasmids using liquid bacterial cultures. Plasmids were then sequenced prior to transfection into eukaryotic cells. Looking forward, transfected cells will be analyzed for mutations.

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CRISPR-Cas9 Induced Mutations of IFI27 Gene

Jereld R. Nicholson Library: Grand Avenue

IFI27 is a gene that has been previously associated with lung clear cell sarcoma as well as Newcastle disease. The importance of the IFI27 gene was explored using mutation inducing procedures involving the CRISPR-Cas9. CRISPR-Cas9 was discovered as defense mechanism in bacterial cells against viral infections. CRISPR-Cas9 is a gene editing technique that is used to cut out a gene of interest, and this is essential in determining how important the IFI27 gene is in cellular function. To target the IFI27 gene, a guide RNA sequence was chosen from important segments of the gene. This guide RNA sequence was inserted into a plasmid called pSpCas9 obtained by MIT. These modified plasmids were purified using antibacterial selection with LB/ampicillin plates. The purified plasmids were characterized using primers and PCR to amplify our desired guide RNA sequence and analyzed using gel electrophoresis to conclude our insertion was successful. Successful insertions promoted the decision to amplify our plasmids using liquid bacterial cultures. Plasmids were then sequenced prior to transfection into eukaryotic cells. Looking forward, transfected cells will be analyzed for mutations.