title: GSM3592414: Kinetin-treated FD fibroblasts 6; Homo sapiens; RNA-Seq
library_construction_protocol: After seven days of treatment, cells were collected, and RNA was extracted using the TRI reagent procedure provided by the manufacturer. RNA-Seq libraries were prepared by the Genomic and Technology Core (GTC) of MGH using strand-specific dUTP method. In brief, RNA sample quality (based on RNA Integrity Number, or RIN) and quantity was determined using the Agilent 2200 TapeStation and between 100-1000 ng of total RNA was used for library preparation. Each RNA sample was spiked with 1 µl of diluted (1:100) External RNA Controls Consortium (ERCC) RNA Spike-In Mix (4456740, ThermoFisher) alternating between mix 1 and mix 2 for each well in the batch. Samples were then enriched for mRNA using polyA capture, followed by stranded reverse transcription and chemical shearing to make appropriate stranded cDNA inserts. Libraries were finished by adding Y-adapters, with sample specific barcodes, followed by between 10-15 rounds of PCR amplification. Libraries were evaluated for final concentration and size distribution by Agilent 2200 TapeStation and/or qPCR, using Library Quantification Kit (KK4854, Kapa Biosystems), and multiplexed by pooling equimolar amounts of each library prior to sequencing.
attributes.tag: GEO Accession
attributes.value: GSM3592414