GSM3382034: Idh1/Pten/p53 - S6; Mus musculus; miRNA-Seq
Total RNA were extracted with miRCURY RNA isolation kit (Exiqon, #300110) according to manufacturer's protocol A total of 500ng total RNA was converted into microRNA NGS libraries using  NEBNEXT library generation kit (New England Biolabs Inc.) according to the  manufacturer's instructions. Each individual RNA sample had adaptors ligated to its  3' and 5' ends and converted into cDNA. Then the cDNA was pre-amplified with  specific primers containing sample specific indexes. After 15 cycle pre-PCR the  libraries were purified on QiaQuick columns and the insert efficiency evaluated by  Bioanalyzer 2100 instrument on high sensitivity DNA chip (Agilent Inc.). The  microRNA cDNA libraries were size fractionated on a LabChip XT (Caliper Inc.) and  a band representing adaptors and 15-40 bp insert excised using the manufacturer's  instructions. Samples were then quantified using qPCR and concentration standards.  Based on quality of the inserts and the concentration measurements the libraries  were pooled in equimolar concentrations (all concentrations of libraries to be pooled  are of the same concentration). The library pool(s) were finally quantified again with  qPCR and optimal concentration of the library pool used to generate the clusters on  the surface of a flowcell before sequencing using v3 sequencing methodology  according to the manufacturer instructions (Illumina Inc.)
GEO
GSM3382034
GEO Accession
GSM3382034