Functional exploration of the yeast (Saccharomyces cerevisiae) genome: use of a mini-Mu transposon to analyse randomly cloned DNA sequences.

The development of mega-sequencing techniques requires new methods for global functional analysis of cloned DNA fragments. We have developed a mini-Mu transposon adapted to yeast cloned DNA fragment analysis. This transposon allows us to do the following in a single construction: (i) to probe yeast cells for the presence of expressed open reading frames (ORFs) in the cloned DNA fragment; (ii) to localize these ORFs in the fragment and determine their transcription orientation; (iii) to use beta-galactosidase protein fusions to study regulation of these ORFs; and (iv) to disrupt the corresponding chromosomal genes. On a 5-kb yeast DNA sequence, we have verified the reliability of this new tool by comparing the data obtained with the mini-Mu transposon to those obtained by classical methods. This transposon should be of immediate use in the yeast genome sequencing programme.