Bovine Genomic DNA Sequencing from Semen

Genomic DNA of an affected bull was prepared from a semen sample following standard protocols using proteinase K digestion and phenol-chloroform extraction. A gDNA sequencing library with 420 bp insert size was prepared using the TruSeq DNA Sample Preparation Kit (Illumina inc., San Diego, CA, USA). The sample was sequenced on an Illumina HiSeq2500 system using TruSeq SBS v3 chemistry (Illumina inc., San Diego, CA, USA) and the 2x100 bp paired-end read module. The fastq-files were generated with the CASAVA bcl2fastq conversion software (version 1.8.3, Illumina inc., San Diego, CA, USA). The alignment of the reads to the University of Maryland reference sequence (UMD3.1, [37 (link)]) was performed with the Burrows-Wheeler Aligner [41 (link)]. The resulting SAM file was converted into a BAM file with SAMtools [42 (link)]. Duplicate reads were identified and marked with the MarkDuplicates command of Picard-tools [43 ].