GSM2460728: smallRNA X^Y (control) testis Replica 2; Drosophila melanogaster; ncRNA-Seq
Total RNA was isolated with Trizol. In order to check the quality of the RNA extracts, samples were run in an 1% TAE-Agarose gel and integrity of the sample was estimated based on the sharpness of the ribosomal RNA bands. Samples that did not meet this quality control were re-extracted. Concentration of all samples was accessed with Qubit 3.0 Fluorometer (Thermo Fisher Scientific). Samples with intact RNA were subjected to library synthesis using the automated Apollo 324™ NGS Library Prep system set to the parameters for smallRNA libraries using the kit PrepX RNA-Seq kit (Wafergen). Final product of each library was subjected to 15 cycles of PCR enrichment following the program suggested by the manufacturer of the kit. Each PCR reaction was carried out with one barcode. Final libraries were purified with magnetic beads and checked with Agilent 2200 TapeStation (Agilent Technologies). Samples were normalized with Qubit 3.0 Fluorometer, and twelve libraries were combined in a single mix. Multiplexed mix was submitted to 50 cycles of single-end sequencing using HiSeq 2500 (Illumina). Two biological replicates were sequenced for each sample.
GEO Accession
GSM2460728