GSM3454754: CapEP_WT_13M_sRNA; Mus musculus; miRNA-Seq
Pooled biological samples instead of biological replicates were used for deep sequencing experiments. Total RNA isolation was performed using Trizol reagent (Invitrogen). Total RNA concentration was measured by Nanodrop ND-2000 (Thermo Scientific), and quality was assessed using an Agilent 2100 Bioanalyzer. For transcriptome sequencing, cDNAs were prepared from full-length mRNA transcripts using a reverse transcriptase with terminal transferase activity. A total of 2.7 μg cDNA for each sample was sonicated to 200 - 400 bp and used for library preparation with the Illumina Nextera DNA Sample Prep Kit following the standard protocol.The libraries were sequenced by Illumina Hiseq 2000. Small RNA sequencing libraries were created following the Illumina TruSeq Small RNA Sample Preparation protocol. In brief, 5' and 3' RNA adapters that were specifically modified to target the ends of small RNA molecules were ligated to 1 μg of high quality total RNA. Reverse transcription was performed to generate cDNA libraries and PCR was used to amplify and add unique index sequences to each library. The amplified cDNA constructs were loaded and separated by polyacrylamide gel electrophoresis (PAGE), and the fraction corresponding to 18 - 40 nt small RNAs was excised, purified and sequenced by Illumina GAIIx orHiseq 2000.
GEO
GSM3454754