title: GSM4555174: Leaf 3, replicate 2, 12 days post inoculation; Fragaria x ananassa; RNA-Seq
library_construction_protocol: Collected leaves were cut into three sections, used as triplicates of 100 mg initial material and extracted in parallel following a modified method of Christou et al. (2014). The extraction buffer (EB) was supplemented with freshly added 2% β-mercaptoethanol (Applichem GmbH, Germany) in order to preserve samples from RNase activity; the powdered leaves were transferred in ice-cold EB and let on ice for 15 min with shaking every 3 min, to allow the extraction buffer to access all plant material and avoid sedimentation of material, instead of directly adding phenol:chloroform:isoamyl alcohol (25:24:1 v/v; AppliChem GmbH, Germany); RNA samples were washed twice with 70% (v/v) ethanol in order to remove traces of phenols and other potentially interfering components; nucleic acid pellet was air-dried at room temperature for 2 min and subsequently dissolved in 30 µL RNase free water on ice for 15 min. Total RNA was treated with DNase I (Macherey-Nagel GmbH & Co., Germany) according to manufacturer's protocol, followed by an ethanol-based RNA precipitation before resuspending the RNA in 30 µl RNase free water. RNA samples were depleted of rRNA with both bacterial and plant Ribo-Zero rRNA Removal Kits (Illumina, San Diego, USA). cDNA libraries were prepared using a TruSeq Stranded mRNA Library Prep kit (Illumina, San Diego, USA). The cDNA libraries were prepared by the Functional Genomics Centre Zurich (University of Zurich, Switzerland).
attributes.tag: GEO Accession
attributes.value: GSM4555174