Truseq sequencing adaptors

For MethylC-seq library generation, genomic DNA was sonicated to an average size of 200 bp using the Covaris sonicator. Sonicated DNA was then purified and end repaired, followed by ligation to methylated Illumina TruSeq sequencing adaptors. Library amplification was performed with KAPA HiFi HotStart Uracil+ DNA polymerase (Kapa Biosystems), using six cycles of amplification. Single-read MethylC-seq libraries (for details, see Supplementary Table 1) were sequenced on the Illumina HiSeq 1500 platform. The sequenced reads in FASTQ format were mapped to in silico bisulfite-converted reference genomes (danRer7, JGI.71 or mm10 for zebrafish, Xenopus and mouse, respectively) using the Bowtie alignment algorithm with the following parameters: -e 120 -l 20 -n 0 (ref. 70 (link)), as previously reported^{71 (link)}. Published paired-read MethylC-seq data^{6 (link)–8 (link)} were mapped with the following parameters: -e 120 -l 20 -n 1 -k 10 -o 4 -I 0 -X 1000. To estimate the bisulfite non-conversion frequency the frequency of all cytosine base calls at reference cytosine positions in the lambda phage genome (unmethylated spike-in control) was normalized by the total number of base calls at reference cytosine positions in the lambda phage genome (Supplementary Table 1).