Nature research paper: A male steroid controls female sexual behaviour in the malaria mosquito
type II was grown in medium containing 2% glucose , 1.7% yeast nitrogen base without amino acids and ammonium sulfate and 5%N ammonium sulfate as the only source of nitrogen. Yeast was recovered by centrifugation and fed ad libitum to mosquito larvae until pupation. Powdered fish food was supplemented to prevent death at the fourth instar stage. Only females were then used in mating experiments with unlabelled males to analyse the male proteome transferred during copulation.
The pellets were resuspended in 50 µl of 0.1% RapiGest in 50 mM ammonium bicarbonate and sonicated in a water bath. Protein concentration was measured for both pellets and supernatants by BCA assay, and the samples were reduced with 5 mM dithiothreiotol alkylated with 15 mM iodoacetamide and digested with trypsin for 1 h at 37 °C . RapiGest was cleaved with the addition of 200 mM HCl followed by 45 min of incubation at 37 °C and centrifugation at 14,000 r.p.m.
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