Intramolecular and intermolecular snRNA cross-links were generated by irradiating HeLa nuclear extract with 365 nm light in the presence of the psoralen derivative AMT. After deproteinization, cross-linked RNAs were resolved by gel electrophoresis and identified as anomalously migrating species by Northern blotting. In addition to the U4/U6 snRNA cross-link, we detected an intermolecular U2/U6 cross-link, as well as several apparently intramolecular U1, U2, and U5 cross-links. Photoreversal of the U2/U6 cross-link with 254 nm irradiation released stoichiometric amounts of U2 and U6 snRNA. To localize the U2/U6 cross-link, the 3' end of U2 in the purified U2/U6 complex was labeled selectively using a novel oligonucleotide "splint" technique. The labeled U2/U6 complex was then subjected to rapid enzymatic RNA sequencing or to targeted digestion of the U2 and U6 components of the complex by RNase H and a panel of complementary oligonucleotides. The U2/U6 cross-link is located upstream of nucleotide 15 in U2 and downstream of nucleotide 85 in U6, suggesting that the phylogenetically conserved base-pairing between these regions (6 consecutive base pairs in human, Drosophila melanogaster, and Caenorhabditis elegans, 7 in Schizosaccharomyces pombe and Trypanasoma brucei, 8 in Pisum sativum, 11 in Saccharomyces cerevisiae) is significant.
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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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