Title

CRISPR-Cas9-Mediated Knockout of Histone Demethylase KDM6A in Pancreatic Cancer

Student Classification

Junior

Faculty Mentor

Dr. Checo Rorie

Department

Biology

Document Type

Poster

Publication Date

Spring 2019

Abstract

Clustered Regularly Interspaced Short Palindromic Repeats(CRISPR)-Cas9 originates from a system in bacteria used to protect them from invading phages and plasmids. CRISPR for genome editing involves the usage of a Cas9 protein and a single guide RNA (sgRNA) that form a complex and target the complementary sequence indicated by the sgRNA. CRISPR-Cas9 genome editing causes double strand breaks and creates random insertions and deletions causing an out of frame transcript. We sought to knockout KDM6A, a histone demethylase, located on the X chromosome. KDM6A is a tumor suppressor gene and is frequently mutated or lost in pancreatic cancer. In pancreatic cancer, KDM6A loss leads to activation of oncogenes, such as ΔNp63, MYC, and RUNX3, due to the deregulation of the COMPASS-like complex. In order to study the molecular mechanisms in human tumorigenesis, we sought to use CRISPR-Cas9 genome editing to knock out KDM6A in difficult to transfect pancreatic cancer cell lines. To achieve this, we will clone into the lentiCRISPR_V2 lentiviral vector to infect the pancreatic cell lines. In pancreatic cancer that retains KDM6A, we aim to determine if CRISPR-mediated knockout of KDM6A causes aberrant activation of oncogenes such as ΔNp63, MYC, and RUNX3, and squamous-like metastatic pancreatic cancer to develop.

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