Improved long-transcript representation in Oxford Nanopore direct RNA sequencing with UltraMarathonRT.

ABSTRACT: While most RNA-seq methods sequence amplified cDNA molecules, the advent of direct RNA sequencing (DRS) empowered the scientific community to read native RNA. This technology unlocked characterization of natural RNA modifications and long RNA isoforms without the inherent biases of PCR amplification. In the library preparation prior to Oxford Nanopore (ONT) sequencing, polyadenylated RNAs are copied by a reverse transcriptase (RT) to generate an RNA-cDNA hybrid. The step aims to eliminate the secondary and tertiary structure inherent to most RNA sequences prior to presentation of the RNA strand to the pore for sequencing. The current recommended protocol for DRS utilizes Induro^® RT and requires reverse transcription at 60°C. We demonstrate that these RT conditions promote hydrol

SUBMITTER: Maio G 

PROVIDER: S-EPMC12713139 | biostudies-literature | 2025 Dec

REPOSITORIES: biostudies-literature