Acute generation of axenic flies without antibiotics

Embryos were bleached and transferred to sterile vials (twenty-five embryos per vial) either without or with antibiotics, respectively. After five days of fly emergence, flies from sterile vials were transferred either to fresh, sterile vials or to vials that had been used to house flies that had emerged from unbleached embryos and raised for an additional five days. Finally, 10-day-old, male flies were frozen and individual vials were examined for presence or absence of bacteria using 16S rDNA PCR (Figure S10). After verification of microbe-free and microbe-contaminated conditions, RNA from head samples was used to perform qPCR using probes listed in Table S15.