MyxoEE-3 culturing protocol
Initiation and transfer protocol
Independent clones of three ancestral motility genotypes (A+S+, A-S+, A+S-) were grown in liquid and used to initiate long-term motility lineages. Eight or twelve replicate populations were established from ancestral clones in each of 16 selective regimes that all shared the same transfer protocol. Each ancestral motility genotype was represented by two genetic marker variants (sensitive vs resistant to rifampicin) and half of the populations within each treatment were founded from each of the two differentially marked ancestor variants. Colonies were established at the center of 15 cm petri dishes and allowed to swarm outward for two weeks at 32 °C and 90% rH. At two-week intervals, an ~3 x 5 mm sample was harvested from the distal-most point on the swarm perimeter and placed in the center of a new plate. This cycle was repeated 40 times for most treatments and 24 times for treatments on TPM agar.
