RNAseq of Mamestra configurata: excised midgut from 10 molting 4th to 5th instar larvae
Midguts were dissected out of 90 molting 5th instar larvae (soon to be 5th instar larvae).  Following removal of the peritrophic matrix and food bolus, the midgut tissue was directly immersed in TRIzol (Life Technologies). The midgut samples in TRIzol were stored at -70oC until tissue collection was complete. Total RNA was extracted by homogenizing in TRIzol reagent according to the manufacturer's protocol (Invitrogen). RNA was resuspended in sterile DEPC-treated H2O and quantified by measuring absorbance at 260 and 280 nm using a Nanodrop spectrophotometer (Fisher-Thermo).  RNA was run through the Micro poly-A purist column (Fisher-Thermo) to select for polyadenylated transcripts. The quality of the RNA was assessed using an RNA 6000 Nano Kit and a Bioanalyzer 2100 instrument (Agilent).454 sequencing libraries were synthesized and sequencing performed at the National Research Council, Saskatoon, Canada