Messenger RNAs transcribed from yeast linear cytoplasmic plasmids possess unconventional 5’ and 3’ UTRs and suggest a novel mechanism of translation

Václav Vopálenský; Michal Sýkora; Tomáš Mašek; Martin Pospíšek

Filter sterilized culture medium was tested for the presence of the killer toxin activity by the agar well diffusion assay using S. cerevisiae S6/1 as a sensitive strain. Approximately 2x10 5 of sensitive yeast cells were plated onto OSS1 agar plates (5.3% worth agar; 0.7% agar; 1% glucose; 1M sorbitol; 0.002% methylene blue; buffered to pH 4.7 with sodium citrate) for testing of M1 killer toxin activity or onto YPD plates (1% yeast extract; 2% peptone; 2% glucose; 2% of agar) for testing of pGKL1 killer toxin activity. Wells were made with an 8 mm diameter cork borer and 100 μl of filter sterilized culture medium or 100 μl of serial dilution of the filter sterilized culture medium was pipetted into well. Inhibition zones were measured after 48 hours of incubation at 24°C.