SeqCapBis sequencing of Q. petraea buds
Custom targeted genome bisulfite sequencing was performed using the SureSelect XT Methyl-Seq Target Enrichment (Agilent, Santa Clara, CA, USA, https://www.agilent.com/en/product/next-generation-sequencing/hybridization-based-next-generation-sequencing-ngs/dna-seq-reagents-kits-library-preparation-kits/sureselectxt-methyl-seq-library-preparation-kit-for-targeted-methylation-sequencing-232864) according to Agilent recommendations. The workflow of the library preparation started with fragmentation to 200 base pairs (bp) of 1.5g of purified DNA, followed by end-repair, A-tailing and ligation of methylated adaptors prior to custom probes (1:8 diluted) hybridization (16h). Bisulfite conversion was then performed with EZ-DNA Methylation-Gold Kit (ZYMO, Irvine, CA, https://www.zymoresearch.com/collections/ez-dna-methylation-gold-kits/products/ez-dna-methylation-gold-kit). 1st PCR was performed with 8 to 9 cycles, 2nd PCR (indexing) was performed with 6 cycles. 96 samples to be sequenced were combined at equimolar concentrations in a single pool and the sequencing was performed in paired ends (2 x 150 pb) on 1 lane of an Illumina NovaSeq6000 S4 flow-cell.