Instrument: Illumina HiSeq 2000
Strategy: RNA-Seq
Source: TRANSCRIPTOMIC
Selection: cDNA
Layout: SINGLE
Construction protocol: 150 mg of seed (based on dry seed weight) was ground and extracted with 1 ml of frozen XT buffer (0.2M sodium borate, 30 mM EGTA, 1% SDS, 1% sodium deoxycholate, 2% polyvinylpyrollidone, 10 mM DTT, and 1% IGEPAL [pH 9.0]) in a pestle and mortar. This was allowed to thaw and was treated with 40 μl proteinase K (PCR grade, Roche, UK) for 90 min at 42°C; precipitation on ice followed for 1 hr with 80 μl 2M potassium chloride. The supernatant was collected after centrifugation at 4°C. The RNA was precipitated from the supernatant at −20°C for 2 hr with 360 μl 8M lithium chloride. The RNA was collected by centrifugation at 4°C and redissolved in 100 μl water. The RNA was further purified via the clean-up protocol of the RNeasy Plant RNA isolation kit (Qiagen) according to the manufacturer's protocol. 5 μg RNA was submitted to sequencing using standard protocols for Illumina TruSeq™ technologies RS-122-2001 | TruSeq™ RNA Sample Prep Kit v2. Data were generated on Illumina HiSeq 2000. The input material contains 1-2% PhiX which is used as a control.