RNA-seq of Col-0 grown at normal temperature
libraries were constructed for 12 RNA samples using the TruSeq RNA protocol (Illumina 15026495 Rev.F). The library preparation involved an initial QC of the RNA using Qubit DNA (Life technologies Q32854) and RNA (Life technologies Q32852) assays as well as a quality check using the Bioanalyser with the Nano kit (Agilent 5067-1511). One g of this RNA was used to obtain mRNA with a poly- A pull down using biotin beads, fragmented at 94C for 6 minutes and random hexamers were used for first-strand cDNA synthesis. The 78 libraries had an average insert size of 270 bp. The insert size of the libraries was verified by running an aliquot of the library on the Agilent bioanalyser using the High Sensitivity chip (Agilent 5067-4626) and the concentration was determined by using a High Sensitivity Qubit assay (ThermoFisher Q32854). A 16-plex equimolar pool was prepared and checked by q-PCR, before preparing and loading for sequencing on the Hiseq 4000 (Illumina) using 150 paired-end reads across 3 lanes. The total number of raw reads generated in the RNA-seq data was ~ 23 M per biological replicate for each sample.
SUB6177755
LIB30105_R2_L8