GSM1666217: testis_6MPN (deg-seq); Mus musculus; OTHER
Tissue total RNA samples for cerebellum and testis were purchased from Zyagen. Total RNA of H1 cells was isolated using Trizol (Life Technologies). Additional column purification and DNaseI treatment were applied using Direct-zol RNA kit (Zymo Research). To generate the degradome sequencing libraries, we used the global 5' RACE library preparation method (Karvinov et al, 2010) with some modifications. Briefly, poly(A)+ mRNA was isolated from 400-500 µg of total RNA using Dynabead Oligo(dT) (Life Technologies) according to the manufacturer's instructions. This procedure was repeated to increase the effectiveness of poly(A) selection. The NEBNext Small RNA library Prep kit was used for 5' adapter ligation, followed by reverse transcription using random hexamer primers containing the 3' SR adapter sequence 5'-AGACGTGTGCTCTTCCGATCTNNNNNN.  PCR was conducted in 25 cycles at 94°C 15s, 60°C 30s, 70°C 1min. The final libraries were purified from 6% PAGE gel followed by AMPure XP Beads size selection (Beckman Coulter).
GEO Accession
GSM1666217