GSM4634678: GM03509 GFP Clone 100; Homo sapiens; miRNA-Seq
RNA was extracted using Trizol and the isolated RNA analyzed s on an Agilent 2100 bioanalyzer for its integrity using the RNA 6000 nano kit (Agilent) to confirm RINs above 8.0. Additionally, the quality of the ribosomal 28 S and 18 S on an agarose gel was used as a measure of integrity and the absorbance at 260/280 nm was used to determine quantity of the extracted RNA. The small RNAs libraries were constructed using Small RNA v1.5 sample preparation kit according to the manufacturer's instructions (Illumina, inc., USA.)  Briefly the total RNA (700-800ng) were ligated to 3′ and 5′ RNA adapters. The ligation products were reverse transcribed using Superscript II Reverse Transcriptase and amplified with 12 cycles of PCR. The PCR products constituting the small RNA cDNA libraries were resolved on 6% Novex TBE PAGE Gel and ~150 bp fragments excised. The library was eluted from the PAGE gel and analyzed on Agilent 2100 Bioanalyzer using DNA high sensitivity kit (Agilent Technologies, USA). Sequencing of miRNA libraries (~150bp fragments) were performed using Illumina GAIIX sequencing platform for 36 cycles.