Application of Long-Read Sequencing to Modified Nucleotides for Detecting Chromatin Accessibility

• Main Author: Saint-John, Brandon
• Format: Dissertation
• Language: English
• Published: ProQuest Dissertations & Theses 01-01-2023
• Subjects: Bioinformatics; Biology; Genetics
• ISBN: 9798380341912

Nucleosomes provide an additional layer of gene regulation by regulating chromatin accessibility. We have found that factors involved in regulating nucleosome positioning, such as SMARCA4, are recurrently mutated in lung adenocarcinoma and can correlate with alternative splicing changes. As a result, it is crucial to understand nucleosome positioning across an entire gene body to understand how they interact to cause changes in splicing. The key to being able to understand this is using long-read sequencing methods to understand the positioning of nucleosomes at once. My thesis focuses on developing methods and computational tools to understand nucleosome positioning with long reads. I developed a sequencing approach called Add-seq that uses a small molecule called angelicin to label accessible regions and determine those positions using nanopore sequencing. Based on the analyses for Add-seq, I also developed a toolkit called cawlr. This computational pipeline can automate calling nucleosomes on single molecules from any long-read sequencing technology. This work provides new ways of looking at nucleosomes and understanding their positioning in the context of other biological processes.