Submission Title

Method for Targeted Disruption of IFIT1

Location

Jereld R. Nicholson Library: Grand Avenue

Subject Area

Biochemistry

Description

IFIT1 is a protein-coding gene known to inhibit viral translation, but the mechanism by which it accomplishes this is unknown. In an effort to understand how IFIT1 works, we’ve designed a system to knock down the gene in human lung cells. Gene knockdown is commonly accomplished using CRISPR-Cas9 technology, which is an endonuclease system that can target and cut specific DNA sequences. The sequence of the short guide RNA (sgRNA), a component of CRISPR-Cas9, is what determines where the endonuclease binds and makes its cut, ultimately resulting in a mutated, non-functional gene. CRISPR-Cas9 is not naturally found in human cells, so to accomplish the gene knockout we had to transfect the cell line with a DNA plasmid encoding the CRISPR complex, including sgRNA complimentary to IFIT1. Our results confirm that we not only successfully inserted our plasmid into the cells, but that we got the plasmid to be expressed as well.

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Method for Targeted Disruption of IFIT1

Jereld R. Nicholson Library: Grand Avenue

IFIT1 is a protein-coding gene known to inhibit viral translation, but the mechanism by which it accomplishes this is unknown. In an effort to understand how IFIT1 works, we’ve designed a system to knock down the gene in human lung cells. Gene knockdown is commonly accomplished using CRISPR-Cas9 technology, which is an endonuclease system that can target and cut specific DNA sequences. The sequence of the short guide RNA (sgRNA), a component of CRISPR-Cas9, is what determines where the endonuclease binds and makes its cut, ultimately resulting in a mutated, non-functional gene. CRISPR-Cas9 is not naturally found in human cells, so to accomplish the gene knockout we had to transfect the cell line with a DNA plasmid encoding the CRISPR complex, including sgRNA complimentary to IFIT1. Our results confirm that we not only successfully inserted our plasmid into the cells, but that we got the plasmid to be expressed as well.