title: GSM3822229: Bull2_WGBS; Bos taurus; Bisulfite-Seq
library_construction_protocol: Sperm DNA was extracted using a proteinase K lysising protocol.  Breifly, frosen semen samples were liquefied at 37°C and were analyzed for concentration, motility and vitality. The semen was centrifuged at 3400 rpm for 5 min to remove supernatant. Then the pellets were washed twice with 1000  µL sterile phosphate buffered saline (PBS, 0.5 M). Then the remaining spermatozoa were incubated overnight in lysis buffer containing dithiothreitol (DDT)and protein K. After lysis, samples were centrifuged to separate the cell debris in the pellet. The supernatant was carefully removed and the sperm DNA was precipitated using ethanol. After that the pellet was dried,  the DNA was redissolved in TE buffer or ddH2O for further use. A total of 5.2 μg of genomic DNA spiked with 26 ng of lambda DNA was fragmented into 200–300 bp, followed by end repair and adenylation. Cytosine-methylated adapters provided by Illumina were ligated to the sonicated DNA. Then, these DNA fragments were treated twice with bisulfite using EZ DNA Methylation-Gold™ kit (Zymo Research), as per the manufacturer's instructions. The resulting DNA fragments were amplified by two cycles of PCR using KAPA HiFi HotStart Uracil + ReadyMix. The libraries were sequenced on an Illumina Hiseq 2500 platform and 125-bp paired-end reads were generated.
attributes.tag: GEO Accession
attributes.value: GSM3822229