Heads were dissected from dcr-2 mutant adult flies (dcr-2^{L811fsX} allele described in Lee et al. (2004) Cell 117(1):69-81) as described in Seitz et al. (Curr. Biol. 18:147-151 (2008)).
Biomaterial provider
Megha Ghildiyal
Extracted molecule
other
Extraction protocol
Total RNA was extracted using the mirVana kit (Ambion), then short (18-30 nt) RNAs were gel-purified. 2S rRNA was depleted as described in Seitz et al. (Curr. Biol. 18:147-151 (2008)).
Label
not applicable
Label protocol
not applicable
Hybridization protocol
not applicable
Scan protocol
not applicable
Description
This RNA sample was beta-eliminated as described in Horwich et al. (2007) Curr. Biol. 17(14):1265-1272 before adapter ligation. This sample was not used in Ghildiyal et al. (2008) Science.
Data processing
Small RNA sequences were extracted from the deep-sequencing reads by the following procedure: For n=29 nt, identify all the reads whose 5´ n-mer is immediately followed by the first trinucleotide (CTG) of the 3´ adapter, and whose 5´ n-mer perfectly maps on the Drosophila melanogaster genome (assembly R5.5 from FlyBase). These inserts are removed from the pool, and flagged as "29 nt-long inserts". Decrease n iteratively (n=28 nt, n=27 nt, ...) and repeat the above procedure at each step, until n=18 nt. Pool all 12 sets (29 nt-long inserts, 28 nt-long inserts, ..., 18 nt-long inserts), to generate the list of genome-matching inserts.
Raw data available at SRA under accession SRX000320
