Recovery of YAC-end sequences through complementation of an Escherichia coli pyrF mutation

We have developed a genetic means to recover sequences from YAC-ends near the yeast selectable marker URA3. This strategy is based on the ability of URA3 to complement mutations in pyrF, an Escherichia coli gene required for pyrimidine biosynthesis. We have developed an E.coli strain with a non-reverting allele of pyrF that is also suitable for cloning (recA-, hsdR-). We demonstrate the utility of this complementation strategy to obtain right-end clones from three YACs containing Arabidopsis thaliana DNA.