Single-step Marker Switching in  Schizosaccharomyces pombe  Using a Lithium Acetate Transformation Protocol.
The ability to utilize different selectable markers for tagging or mutating multiple genes in  Schizosaccharomyces pombe  is hampered by the historical use of only two selectable markers,  ura4 +   and  kanMX6 ; the latter conferring resistance to the antibiotic G418 (geneticin). More markers have been described recently, but introducing these into yeast cells often requires strain construction from scratch. To overcome this problem we and other groups have created transformation cassettes with flanking homologies to  ura4  +  and  kanMX6  which enable an efficient and time-saving way to exchange markers in existing mutated or tagged fission yeast strains. Here, we present a protocol for single-step marker switching by lithium acetate transformation in fission yeast,  Schizosaccharomyces pombe . In the following we describe how to swap the  ura4 +   marker to a  kanMX6, natMX4,  or  hphMX4  marker, which provide resistance against the antibiotics G418, nourseothricin (clonNAT) or hygromycin B, respectively. We also detail how to exchange any of the  MX  markers for nutritional markers, such as  arg3 + , his3 + , leu1 +   and  ura4 +   .
Brown SD, Lorenz A