Conventional isolation and detection methods for small RNAs from yeast cells have been designed for a limited number of samples. In order to be able to conduct a genome-wide assessment of how each gene product impacts upon small RNAs, we developed a rapid method for analysing small RNAs from Saccharomyces cerevisiae wild-type (wt) and mutants cells in the deletion and temperature-sensitive (ts) collections. Our method implements three optimized techniques: a procedure for growing small yeast cultures in 96-deepwell plates, a fast procedure for small RNA isolation from the plates, and a sensitive non-radioactive northern method for RNA detection. The RNA isolation procedure requires only 4 h for processing 96 samples, is highly reproducible and yields RNA of good quality and quantity. The non-radioactive northern method employs digoxigenin (DIG)-labelled DNA probes and chemiluminescence. It detects femtomole levels of small RNAs within 1 min exposure time. We minimized the processing time for large-scale analysis and optimized the stripping and reprobing procedures for analyses of multiple RNAs from a single membrane. The method described is rapid, sensitive, safe and cost-effective for genome-wide screens of novel genes involved in the biogenesis, subcellular trafficking and stability of small RNAs. Moreover, it will be useful to educational laboratory class venues and to research institutions with limited access to radioisotopes or robots.
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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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Increase the total number of rows showing on this page by using the pull-down located below the table, or use the page scroll at the table's top right to browse through the table's pages; use the arrows to the right of a column header to sort by that column; filter the table using the "Filter" box at the top of the table; click on the small "i" buttons located within a cell for an annotation to view further details about experiment type and any other genes involved in the interaction.
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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