GSM2213647: GQF-6; Mus musculus; miRNA-Seq
Mouse hippocampal tissue was homogenized using TissueLyser (Qiagen) in Trizol® reagent (Invitrogen). Total RNA was extracted by phenol-chlorofom precipitation. The quality and quantity of RNA was determined by running the samples on a Bioanalyzer ® (Agilent) and Qubit® fluorometer (Invitrogen) respectively. Small RNA libraries were prepared from 1ug of total RNA using TruSeq® Small RNA Kit (Illumina) according to manufacturer’s instructions. Briefly, 3’ and 5’ adapters were ligated to small RNAs using T4 DNA ligase. Ligated RNA was reverse transcribed to cDNA using superscript II reverse transcriptase. The resulting cDNA template was amplified by PCR to generate a cDNA library. For each sample, a common forward primer (which binds to the 5’ adapter complement) and a unique reverse primer (which binds to the 3’ adapter complement and contains a unique sequence for each sample) were used.  The quality of the resulting amplicons was analyzed on a high sensitivity DNA ChIP (Bioanalyzer). Next, the cDNA construct was run on a 6% PAGE gel and amplicons corresponding to adapter ligated miRNA sequences (145-160nt) were excised out. DNA from excised gel was eluted and precipitated in Ethanol. The size, purity, and amount of cDNA was assessed on high sensitivity DNA ChIP (Bioanalzyer).  Then a titration run was done to check quality of the library and validate the amount.
GEO Accession
GSM2213647