Project description:RNA internal modifications play critical role in development of multicellular organisms and their response to environmental cues. Using nanopore direct RNA sequencing (DRS), we constructed a large in vitro epitranscriptome (IVET) resource from plant cDNA library labeled with m6A, m1A and m5C respectively. Furthermore, after transfer learning, the pre-trained model was used to detect additional RNA internal modification such as m1A, hm5C, m7G and Ψ modification. Finally, we illustrated a global view of epitranscriptome with m6A, m1A, m5C, m7G and Ψ modification in rice seedlings under normal and high salinity environment. In summary, we provided a strategy for creating IVET resource from cDNA library and developed a computational method that use IVET-based transfer learning termed TandemMod for profiling epitranscriptome landscape with co-occupancy of multiple types of RNA modification in plants responsive to environmental signal.
Project description:5-methylcytosine (5mC) is an important type of epigenetic modification. In this study, we enhanced 5mC detection using SMRT sequencing by holistically analyzing kinetic signals of a DNA polymerase and sequence context for every base within a measurement window. We employed a convolutional neural network to train a methylation classification model. This methodology has provided a system for simultaneous genome-wide genetic and epigenetic analyses.ÂÂ
Project description:Cancer is a complex disease, driven by a combination of genetic and epigenetic alterations. DNA and RNA methylation modifications are the most common epigenetic events that play critical roles in cancer development and progression. Bisulfite converted sequencing is a widely used technique to detect base modifications in DNA methylation, but its main drawbacks lie in DNA degradation, lack of specificity, or short reads with low sequence diversity. The nanopore sequencing technology can directly detect base modifications in native DNA as well as RNA without harsh chemical treatment, compared to bisulfite sequencing. Furthermore, CRISPR/Cas9-targeted enrichment nanopore sequencing techniques are straightforward and cost-effective when targeting genomic regions are of interest. In this review, we mainly focus on DNA and RNA methylation modification detection in cancer with the current nanopore sequencing approaches. We also present the respective strengths, weaknesses of nanopore sequencing techniques, and their future translational applications in identification of epigenetic biomarkers for cancer detection and prognosis.
Project description:The covalent modification of RNA molecules is a pervasive feature of all classes of RNAs and has fundamental roles in the regulation of several cellular processes. Mapping the location of RNA modifications transcriptome-wide is key to unveiling their role and dynamic behaviour, but technical limitations have often hampered these efforts. Nanopore direct RNA sequencing is a third-generation sequencing technology that allows the sequencing of native RNA molecules, thus providing a direct way to detect modifications at single-molecule resolution. Despite recent advances, the analysis of nanopore sequencing data for RNA modification detection is still a complex task that presents many challenges. Many works have addressed this task using different approaches, resulting in a large number of tools with different features and performances. Here we review the diverse approaches proposed so far and outline the principles underlying currently available algorithms.
Project description:Pseudouridine (Ψ) is an abundant mRNA modification in the mammalian transcriptome, but its function has remained elusive due to the difficulty of transcriptome-wide mapping. We develop nanopore native RNA sequencing for quantitative Ψ analysis that utilizes native content training, machine learning model prediction, and single read coordination. We find interferon inducible Ψ modifications in the interferon stimulated gene transcripts, consistent with a role of Ψ in the efficacy of mRNA vaccines.