ATACseq of liver, muscle and hypothalamus of three Brahman heifers

Liver, hypothalamus, and muscle samples were collected from three unrelated, post-pubertal Brahman heifers of similar age and weight. ATAC-seq libraries were prepared from frozen tissues and sequenced on an Illumina NextSeq 150 cycle (2X 75 bp). Data was used to identify tissue-specific open chromatin regions in indicine cattle and identify master regulators (transcriptions factors) of each tissue.\nLineage: Liver, hypothalamus and muscle samples were collected from three unrelated, post-pubertal Brahman heifers of similar age and weight. Heifers used in this study were managed, handled, and euthanized as per approval of the Animal Ethics Committee of the University of Queensland, Production and Companion Animal group (certificate number QAAFI/279/12). After slaughter, tissue samples were collected as fast as possible and stored at−80◦C. ATAC-seq libraries were prepared from frozen tissues using the Omni-ATAC method with the following modifications. Frozen tissue (20mg) was ground in liquid nitrogen using a mortar and pestle. The pulverized tissue was transferred to a pre-chilled 2ml Dounce homogenizer containing 1ml cold 1 x homogenization buffer and homogenized with the pestle until a uniform suspension was seen (10-20 strokes). The homogenate was filtered with a 40uM nylon cell strainer (BD Falcon) before layering onto the iodixanol solution as described previously. The ratio of nuclei to enzyme concentration was optimized for each sample by performing transposition reactions containing 50 000, 100 000 and 200 000 nuclei with 2.5ul of tagment enzyme in 50ul of transposition mix. The transposed DNA was amplified with custom primers. Amplified libraries were purified using Agencourt AMPure XP beads (Beckman Coulter) and quality controlled using a Bioanalyser High Sensitivity DNA Analysis kit (Agilent). ATAC-seq libraries were sequenced at IMB sequencing facility (University of Queensland) on an Illumina NextSeq 150 cycle (2X 75 bp). Three biological replicates were performed per tissue. All assays were performed according to FAANG guidelines and recommendations, available at http://www.faang.org.