GSM3133608: Control Total Small RNA; Drosophila melanogaster; ncRNA-Seq
Thirty μg of total RNA from Control (mat>Luciferase) and Nup358 GLKD ovaries was resolved on a 15% denaturing polyacrylamide gel containing 7M urea. Small RNAs ranging from 20-40nt were purified without oxidation step as described in Li C et al (Cell, 137, 509-521). Small RNA sequencing libraries were prepared  using  TruSeq Small RNA Library Preparation Kit (Illumina, San Diego, CA) with one minor modification. A terminator block oligo (5'-TAC AAC CCT CAA CCA TAT GTA GTC CAA GCA/3SpC3/-3') was added prior to 5' adapter ligation as described in Wickersheim, M.L et al (Biotechniques,55,269-272). This step specifically blocked reverse transcription of 30nt long 2S rRNA and hence substantially reduced rRNA contamination in the final sequenced reads.   Sequencing libraries were size selected with a Pippin Prep (Sage Science, MA). Single read sequencing (1X50 cycles) was performed on a HiSeq2500 (Illumina, San Diego, CA). Sequencing was performed at Hollings Cancer Center Genomics Core Lab at Medical University of South Carolina (MUSC).