The Complete Set of Predicted Genes from Saccharomyces cerevisiae in a Readily Usable Form.
Hudson JR Jr, Dawson EP, Rushing KL, Jackson CH, Lockshon D, Conover D, Lanciault C, Harris JR, Simmons SJ, Rothstein R, Fields S
Genome Research (1997)
Category: methods ¤ Added: Jan 30, 2003 ¤ Rating: ◊◊
Nearly all of the open reading frames (ORFs) of the yeast Saccharomyces cerevisiae have been synthesized by PCR using a set of ~6000 primer pairs. Each of the forward primers has a common 22-base sequence at its 58 end, and each of the back primers has a common 20-base sequence at its 58 end. These common termini allow reamplification of the entire set of original PCR products using a single pair of longer primers—in our case, 70 bases. The resulting 70-base elements that flank each ORF can be used for rapid and efficient cloning into a linearized yeast vector that contains these same elements at its termini. This cloning by genetic recombination obviates the need for ligations or bacterial manipulations and should permit convenient global approaches to gene function that require the assay of each putative yeast gene.