GSM1874041: S17_age16.93; Macaca mulatta; RNA-Seq
Tissues were homogenized in Trizol (Invitrogen, NY) and used to isolate total RNA according to the manufacturer's protocol.  Total RNA (5 μg) was treated with 20 U of RNase R (RNR07250, Epicenter) for 15 min at 37⁰C and the digested RNA was isolated using acidic phenol-chloroform (5:1) and ethanol precipitation.  CircRNA sequencing was performed at the Genomics Research Center, University of Rochester; 24 samples (one per monkey) were sequenced initially at a depth of approximately 20 million reads (paired-end), and two samples were subsequently sequenced in-depth (at approximately 100 million reads) to assess sample saturation. Modified TruSeq RNA.  Briefly, quantitation and quality of RNA was determing with NanoDrop and Agilent Bioanalyzer 2100, respectively.  PolyA capture with oligo-dT beads was omitted and each library was generated from 40ng RNA input with a fragmentation time of 5 minutes at 94C. First-strand cDNA synthesis was performed with random hexamer priming followed by second-strand cDNA synthesis. End repair and 3` adenylation was then performed on the double stranded cDNA. Illumina adaptors were ligated to both ends of the cDNA, purified by gel electrophoresis and amplified with PCR primers specific to the adaptor sequences to generate amplicons of approximately 200-400bp in size. The amplified libraries were hybridized to the Illumina pair end flow cell and amplified using the cBot
GEO Accession
GSM1874041