Bacteria and yeast colony PCR

Abstract

The bacteria Escherichia coli and the yeast Saccharomyces cerevisiae are currently the two most important organisms in synthetic biology. E. coli is almost always used for fundamental DNA manipulation, while yeast is the simplest host system for studying eukaryotic gene expression and performing large-scale DNA assembly. Yeast expression studies may also require altering the chromosomal DNA by homologous recombination. All these studies require the verification of the expected DNA sequence, and the fastest method of screening is colony PCR, which is direct PCR of DNA in cells without prior DNA purification. Colony PCR is hampered by the difficulty of releasing DNA into the PCR mix and by the presence of PCR inhibitors. We hereby present one protocol for E. coli and two protocols for S. cerevisiae differing in efficiency and complexity as well as an overview of past and possible future developments of efficient S. cerevisiae colony PCR protocols.