In addition to A, C, G and U, RNA contains over 100 additional chemically distinct residues. An abundant modified base frequently found in tRNAs, dihydrouridine (D) has recently been mapped to over 100 positions in mRNAs in yeast and human cells. Multiple highly conserved dihydrouridine synthases associate with and modify mRNA, suggesting there are many D sites yet to be found. Because D alters RNA structure, installation of D in mRNA is likely to effect multiple steps in mRNA metabolism including processing, trafficking, translation, and degradation. Here, we introduce D-seq, a method to chart the D landscape at single nucleotide resolution. The included protocols start with RNA isolation and carry through D-seq library preparation and data analysis. While the protocols below are tailored to map Ds in mRNA, the D-seq method is generalizable to any RNA type of interest, including non-coding RNAs, which have also recently been identified as dihydrouridine synthase targets.
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| Evidence ID | Analyze ID | Gene/Complex | Systematic Name/Complex Accession | Qualifier | Gene Ontology Term ID | Gene Ontology Term | Aspect | Annotation Extension | Evidence | Method | Source | Assigned On | Reference |
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| Evidence ID | Analyze ID | Gene | Gene Systematic Name | Phenotype | Experiment Type | Experiment Type Category | Mutant Information | Strain Background | Chemical | Details | Reference |
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| Evidence ID | Analyze ID | Gene | Gene Systematic Name | Disease Ontology Term | Disease Ontology Term ID | Qualifier | Evidence | Method | Source | Assigned On | Reference |
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| Evidence ID | Analyze ID | Regulator | Regulator Systematic Name | Target | Target Systematic Name | Direction | Regulation of | Happens During | Regulator Type | Direction | Regulation Of | Happens During | Method | Evidence | Strain Background | Reference |
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| Site | Modification | Modifier | Source | Reference |
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| Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Allele | Assay | Annotation | Action | Phenotype | SGA score | P-value | Source | Reference | Note |
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| Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Assay | Annotation | Action | Modification | Source | Reference | Note |
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| Complement ID | Locus ID | Gene | Species | Gene ID | Strain background | Direction | Details | Source | Reference |
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| Evidence ID | Analyze ID | Dataset | Description | Keywords | Number of Conditions | Reference |
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