Grant: Undergraduate Student Research Awards (USRA)

Circulating tumor DNA (ctDNA) consists of short DNA fragments released from tumor cells into extracellular environments during processes such as apoptosis and necrosis. Quantifying ctDNA dynamics provides valuable insights into tumor cell turnover and DNA stability. This project aims to characterize the release and degradation dynamics of ctDNA using a colorectal cancer cell culture model. The HCT116 human colorectal carcinoma cell line will be used to establish baseline parameters governing ctDNA production and detection in vitro. Cells will be cultured under controlled conditions and exposed to X-ray radiation to stimulate cellular stress and DNA release. Cell culture supernatant will be collected at defined time points before and after radiation exposure to monitor ctDNA release dynamics. Cell-free DNA will be isolated from the supernatant using circulating nucleic acid extraction methods optimized for short DNA fragments. Extracted ctDNA will then undergo library preparation for next-generation sequencing using standardized adapter ligation and amplification protocols. Sequencing will be performed using high-throughput paired-end sequencing to identify ctDNA fragments and quantify mutation frequencies associated with the HCT116 genome. Bioinformatics analysis will determine ctDNA release rates, degradation kinetics, and limits of detection within the culture system. The outcomes of this project will establish baseline parameters for ctDNA detection and provide fundamental insights into ctDNA stability and quantification in colorectal cancer cell models, supporting the development of sequencing-based approaches for studying tumor-derived DNA.