Elli Gasper
Pronouns:
Research Mentor(s): Adelaide Tovar
Research Mentor School/College/Department: Computational Medicine & Bioinformatics / Medicine
Program: UROPF
Session: Session 7 (4:40pm – 5:30pm)
Authors: Elli Gasper, Adelaide Tovar, Jacob Kitzman, Stephen Parker
Abstract
The purpose of this project is to assess the regulatory activity of diabetes-associated genetic variants with the use of massively parallel reporter assays (MPRAs) and chromatin profiling. This research will take place in a rat beta cell tissue culture and includes developing an assay to look at changes in chromatin availability for an integrated MPRA. The goal of this research is to determine the chromatin accessibility in DNA around sites where a diabetes-associated genetic variant has been integrated as part of a synthetic gene expression construct. By comparing pairs of alleles for different variants, we will learn more about how changes in gene regulation may lead to diabetes. To complete this project we will perform assay for transposase-accessible chromatin using sequencing (ATAC-seq) in the 832/13 rat insulinoma (beta) cell line. ATAC-seq consists of isolating the nuclei from cells, and therefore being able to use Tn5 transposase to selectively cut DNA where it’s open and no histones are present. Tn5 is special because in addition to cutting, it also adds an adaptor to every piece of DNA allowing for PCR to occur. After doing so, a post-transposition enrichment step occurs to direct PCR primers targeted around the flanking region, oligo and barcode. One set of the 832/13 cells is infected with an insulin promoter, whereas the other is infected with a minimal synthetic promoter. By using the ATAC-seq it will show where the virus has integrated in the genome. With that, we will be able to see whether differences in chromatin accessibility occur across alleles at diabetes-associated genetic variants. This will enable future studies to determine how changes in gene regulation lead to diabetes.
