Exposure of Control and Pesticide-selected Populations to Pesticide and Bt Oral Diets
After eight generations of selection, we generated same-age F2 larvae from pesticide-selected and susceptible colonies. Approximately 200 adults from Gen. 8 pesticide-selected and susceptible control colonies were allowed to reproduce on control diet for three days. We removed these F0 adults and allowed offspring to develop to F1 adults. To generate F2 experimental larvae, we placed 4-5 replicates of 80 adults per selection line in 100 mm petri dishes with control diet and allowed them to reproduce for 24 hours. F2 larvae were allowed to develop for 12 days.
We placed twelve day old larvae (~4 mm long) on oral diet plates prepared as described in the previous section (Suppl. Fig. 1; n = 96/ population/ treatment). Half of the plate (n = 48) was monitored daily for three weeks for survival and development. Individuals in the remaining half were sampled after two days of exposure prior to the onset of mortality and used for RNAseq analyses (described below).