methylC-seq of Arabidopsis rosette: RRGD RRGD Timepoint III - excess-light 60 min rep  5
DNA was extracted using the Qiagen DNeasy Plant Mini Kit as per the manufactureres instructions. In total 1000 ng of gDNA was fragmented using a Covaris (Covaris), column purified using the MinElute PCR Cleanup (Qiagen), end repaired using the End-It Kit (Epicentre), A-tailed then bead cleaned using AMPure XP beads (Beckman Coulter). NEXTflex methylated adapters (Bioo Scientific) were ligated and following bead clean-up the ligation product was quantified on a Qubit (Life Technologies). 450 ng of clean ligation was then bisulfite (BS) converted using the MethylCode Kit (Life Technologies). BS-converted DNA was amplified with 6 cycles of PCR using a Kapa HiFi HotStart U+ 2x readymix (Kapa Biosystems) and an Illumina compatible NEXTflex Primer mix (Bioo Scientific). Samples were bead-cleaned, quantified by qPCR using a Library Quantification Kit for Illumina/LightCycler 480 (Kapa Biosystems), pooled equal-molar and 6-sample pools were sequenced per lane on a Hiseq2500 (100bp single end) .