title: GSM1262122: RNA-seq_WT-2; Mus musculus; RNA-Seq
library_construction_protocol: Dentate gyri were rapidly micro-dissected bilaterally from adult mice. Total RNA was extracted using the RNeasy kit (Qiagen) with on-column DNaseI-treatment. Poly(A)-containing mRNA was purified from 1 µg DNase-treated total RNA. Following the purification, the mRNA was fragmented into small pieces using divalent cations under elevated temperature. Reverse transcriptase and random primers were used to generate the first-strand cDNA. The second-strand cDNA was synthesized using DNA polymerase I and RNaseH. These cDNA fragments went through an end-repair process using T4 DNA polymerase, T4 PNK and Klenow DNA polymerase, and the addition of a single ‘A’ base using Klenow Fragment (3' to 5' exo minus), then the ligation of the Illumina PE adapters using T4 DNA ligase. An index (up to 12) was inserted into Illumina adapters so that multiple samples can be sequenced in one lane of 8-lane flow cell if necessary. These products were then purified and enriched with PCR to create the final cDNA library for Illumina HiSeq2000 sequencing.
attributes.tag: GEO Accession
attributes.value: GSM1262122