RRBS of F2 Parus major
High-quality genomic DNA (1 _g) was sent to the Carver High-Throughput DNA Sequencing and Genotyping Unit (University of Illinois, Urbana, IL, USA) for generation of reduced representation bisulfite sequencing (RRBS) libraries following standard protocols. The final libraries were quantified using Qubit (Life Technologies, Carlsbad, CA, USA) and the average size was determined on an Agilent bioanalyzer DNA7500 DNA chip (Agilent Technologies, Wilmington, DE, USA) and diluted to 10 nM. The 10 nM dilution was further quantitated by qPCR on an ABI 1900 to ensure high accuracy quantification for consistent pooling of barcoded libraries and maximization of the number of clusters in the Illumina flowcell. RRBS Illumina sequencing was performed on multiplexed libraries and loaded onto 8-lane flowcells for cluster formation and sequenced on an Illumina HiSeq2500. The libraries were sequenced to a total read length of 100 bp from one end (single-end sequencing) of the molecules. Fastq files were generated and demultiplexed with with the bcl2fastq v2.17.1.14 Conversion Software (Illumina, San Diego, CA, USA).