GSM1313428: AA-treated_2; Rattus norvegicus; RNA-Seq
40-50 mg tissue was cut and mechanically minced using Tissue Tearor (Biospec Products Inc, OK).  Total RNA was isolated using a small RNA-retaining protocol that employed an organic extraction followed by glass-fiber immobilization (Ambion, TX).  RNA concentration was determined by Nandrop1000 spectrophotometer (Thermo Scientific, DE). Samples were prepared using illumina small RNA sample Prep kit with the small RNA v1.5 sample preparation guide.  Approximately 10 µg of total RNA was used for small RNA library construction. The v1.5 sRNA 3’ and SRA 5’ adaptors (Illumina, USA) were added to both ends of the small RNA. The 3’ and 5’ Ligated RNAs were used as templates for reverse transcription followed by PCR amplification.  The enriched cDNA constructs were loaded to 6% TBE PAGE gel and the bands containing the 22-30 nt RNA fragments (93-100 nt in length with both adapters) were cut and purified. The concentrations of the libraries were determined using a NanoDrop ND-1000 Spectrophotometer and the size and purity were determined using an Agilent 2100 Bioanalyzer in combination with the Agilent DNA 1000 Kit. The purified DNA was used directly for cluster generation and sequence analysis using the Illumina Genome Analyzer II (Illumina, San Diego, CA, USA) according to the manufacturer's instructions (36 cycle single read cluster kit v4 and sequence kit v4).
GEO Accession
GSM1313428