Presentation description

Oncogenic transcription factors (TFs) are TFs that become hyper-activated in cancer cells and are known to facilitate gene expression. They are crucial to understanding cancer development and can help identify novel therapeutic opportunities. One method of learning more about such oncogenic TFs is Chromatin Immunoprecipitation coupled to sequencing (ChIP-Seq), which is used to identify DNA binding sites of such TFs, allowing us to map them across the genome. Identifying where TFs bind is important for understanding which genes are regulated by the respective TF. An example of an oncogenic TF is SOX2. SOX2 has been previously implicated in squamous cell carcinoma and glioblastoma. We performed ChIP-seq of SOX2 within two cell lines, EPC2, which are esophageal epithelial cells and are the non-cancerous counterpart to squamous cell carcinoma, and U251, which originates from glioblastoma tumors. We performed ChIP-qPCR to assess the specificity of the IP before high-throughput sequencing. We used primers against positive and negative control regions. Positive control regions are regions that are known to be bound by SOX2 and we expect will be enriched if our ChIP works successfully, whereas we do not expect any specific enrichment of negative control regions. We also tested two experimental regions in which we do not know if SOX2 is binding. Through our ChIP-qPCR analysis, we found that the U251 cell line ChIP worked, while the EPC2 cell line ChIP did not. We plan on repeating the EPC2 cell line's ChIP in the future. Additionally, I performed molecular cloning to create plasmids that will be used for various CRISPRi and CRISPRa experiments.