The 2 mu plasmid of the yeast Saccharomyces cerevisiae encodes a site-specific recombination system consisting of the FLP protein and two inverted recombination sites on the plasmid. The minimal fully functional substrate for in-vitro recombination in this system consists of two FLP protein binding sites separated by an eight base-pair spacer sequence. We have used site-directed mutagenesis to generate every possible mutation (36 in all) within 11 base-pairs of one FLP protein binding site and the base-pair immediately flanking it. The base-pairs within the binding site can be separated into three classes on the basis of these results. Thirty of the 36 sequence changes, including all three at seven different positions (class I) produce a negligible or modest effect on FLP protein-promoted recombination. In particular, most transition mutations are well-tolerated in this system. In only one case do all three possible mutations produce large effects (class II). At three positions, clustered near the site at which DNA is cleaved by FLP protein, one of the two possible transversions produces a large effect on recombination, while the other two changes produce modest effects (class III). For seven mutants for which FLP protein binding was measured, a direct correlation between decreases in recombination activity and in binding was observed. Positive effects on the reaction potential of mutant sites are observed when the other FLP binding site in a single recombination site is unaltered or when the second recombination site in a reaction is wild-type. This suggests a functional interaction between FLP binding sites both in cis and in trans. When two mutant recombination sites (each with 1 altered FLP binding site) are recombined, the relative orientation of the mutations (parallel or antiparallel) has no effect on the result. These results provide an extensive substrate catalog to complement future studies in this system.
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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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