Detection of genetic alteration events by ionizing radiation in human embryonic stem cells via next-generation sequencing

Friday, November 08, 2013 — Poster Session III
10:00 a.m. – 12:00 p.m.	FAES Academic Center (Upper-Level Terrace)	CC	STEMCELL-11

Authors

V. Nguyen
I.G. Panyutin
I.V. Panyutin
R.D. Neumann

Abstract

Ionizing radiation (IR) is widely employed for medical purposes from therapy to diagnosis. Although its cytotoxic properties have long been successfully exploited for therapeutics, the molecular mechanism underlying its effects from clinically relevant diagnostic doses remains unclear. We therefore seek to investigate this urgent question as IR-related diagnostic test uses continue to rise. We established a novel model based on human embryonic stem cell (hESC) culture to conduct the study. Cells were subjected to IR at low, diagnostic equivalent (0.2 Gy) or high (1 Gy) doses then maintained in culture until confluence before harvested for DNA isolation and sequencing. As IR is often implicated as a risk for cancer, a primer pool targeting genomic “hot spot” regions that are frequently mutated in human cancer genes was used to generate libraries from low-, high-dose and control samples of H1 hESC line. Utilizing a semiconductor-based sequencing approach, we detected an additional point-mutation event on the KIT proto-oncogene in the high-dose sample but not in the low-dose compared to control. We are expanding the study to other three commonly used hESC lines and plan to confirm our results at the transcription level with RNA sequencing panels.