University of Connecticut OpenCommons@UConn Doctoral Dissertations University of Connecticut Graduate School 6-7-2018 Development and Application of Ribose Oxidation Sequencing (RibOxi-seq) Yinzhou Zhu University of Connecticut,
[email protected] Follow this and additional works at: https://opencommons.uconn.edu/dissertations Recommended Citation Zhu, Yinzhou, "Development and Application of Ribose Oxidation Sequencing (RibOxi-seq)" (2018). Doctoral Dissertations. 1881. https://opencommons.uconn.edu/dissertations/1881 Development and Application of Ribose Oxidation Sequencing (RibOxi-seq) Yinzhou Zhu, PhD University of Connecticut, 2018 In eukaryotes, a number of RNA sites are modified by 2’-O methylation (2’-OMe). Such editing is mostly guided by BoxC/D class small nucleolar RNAs (snoRNAs). These snoRNAs direct methylation via complementary RNA-RNA interactions. 2’-OMe has so far been shown to be present in rRNAs, tRNAs and some small RNAs and has been implicated in ribosome maturation and translational circuitries. A substantial portion of known methylated sites in rRNA lie in close proximity to ribosome functional sites such as regions around the peptidyl transferase center. It is not yet clear whether many mRNAs might possess internal 2’-OMe sites. It is therefore important to characterize 2’-OMe landscapes. We have developed a novel method for the highly accurate and transcriptome-wide detection of 2’-OMe sites. The core principle of this method is to randomly digest RNAs to expose 2’-OMe sites at the 3’-ends of digested RNA fragments. Next, an oxidation step using sodium periodate destroys all fragment-3’-ends except those that are 2’-O methylated. Only these oxidation-resistant fragments are available for linker ligation and subsequent sequencing library preparation.