A yeast null mutant (erg 3) defective in ERG 3, the gene encoding the C-5 sterol desaturase required for ergosterol synthesis was transformed with an Arabidopsis thaliana cDNA library inserted in a yeast vector. Transformants (4 x 10(5)) were screened for cycloheximide (CH) resistance and 400 possible clones were analyzed to determine their sterol profile. Low levels of ergosterol in addition to delta 7- and delta 8-sterols normally present in erg3 were isolated in three yeast transformants. Characterization of one transformant indicated a cDNA of 1141 bp. Transformation of an erg 3 strain with this plasmid led to CH resistance, nystatin sensitivity and an ergosterol profile. After subcloning in a pBluescript vector and subsequent sequencing, an ORF of 843 bp encoding a possible 281 amino acid polypeptide was deduced. Three histidine-rich motifs (HX3H, HX2HH and HX2HH) were found in the A. thaliana ORF which are also present in the yeast ERG 3 gene. These histidine-rich motifs are also characteristic of many membrane-bound fatty acid desaturases from higher plants. These data strongly suggest that the A. thaliana cDNA encodes a delta 7-sterol-C-5-desaturase.
|Evidence ID||Analyze ID||Interactor||Interactor Systematic Name||Interactor||Interactor Systematic Name||Type||Assay||Annotation||Action||Modification||Phenotype||Source||Reference||Note|
|Evidence ID||Analyze ID||Gene||Gene Systematic Name||Gene Ontology Term||Gene Ontology Term ID||Qualifier||Aspect||Method||Evidence||Source||Assigned On||Reference||Annotation Extension|
|Evidence ID||Analyze ID||Gene||Gene Systematic Name||Phenotype||Experiment Type||Experiment Type Category||Mutant Information||Strain Background||Chemical||Details||Reference|
|Evidence ID||Analyze ID||Regulator||Regulator Systematic Name||Target||Target Systematic Name||Experiment||Conditions||Strain||Source||Reference|