Ryan Shafi
Pronouns: he/him
Research Mentor(s): Xin Tong
Co-Presenter: Josyabhatla, Mahadev
Research Mentor School/College/Department: Molecular & Integrative Physiology / Medicine
Presentation Date: April 20
Presentation Type: Poster
Session: Session 6 – 4:40pm – 5:30 pm
Room: League Ballroom
Authors: Ryan Shafi, Xin (Tony) Tong, Rafee Mirza, Mahadev Josyabhatla
Presenter: 120
Abstract
Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) is a technology used to edit genomes at very high precision. It enables precise editing of genomic loci with a RNA-guided Cas9 nuclease that can cleave target DNA that is complementary to a guide RNA (gRNA). Then, the CRISPR-Cas9 system can be used to achieve various goals such as treating inherited diseases (e.g., cystic fibrosis). The objective is to test the specificity and function of the CRISPR-Cas9 system in deleting a targeted sequence in mammalian cells. Mammalian 293AD cells were first transfected with the Cas9-expressing plasmid and human EMX2 ()-specific 3.1+4.1 gRNA sequences. 72 hours after transfection, the DNA will be extracted and purified from the cells. Since the sequence which encodes for green fluorescent protein (GFP) is inside the inserted plasmid, transfection success will be confirmed by the presence of green-glowing cells when imaged. The upcoming steps involve amplifying the hEMX2 via polymerase chain reaction (PCR) and running on a 2% agarose gel to confirm the size and location of DNA fragments when compared to a marker. It is predicted that the gel will display a combination of both WT and Deletion allele. Long-term, the objective of this study is to translate the CRISPR-Cas9 system to hepatic cells and eventually study them in vivo with mice to further understand non-alcoholic fatty liver disease.
Interdisciplinary, Natural/Life Sciences
