GSM3723830: Adult male hypoxia RRBS 3; Rattus norvegicus; Bisulfite-Seq
For genomic DNA (gDNA) extraction, heart tissues were minced and digested with 20 μL proteinase K (20 mg/mL) in digestion buffer (10mM Tris-Hcl, pH 8.0 containing 0.5 M EDTA, 1% SDS, 10 m NaCl, and 5mM CaCl2) for 3 h at 55°C. Next the tissue lysate was twice phenol/chloroform/isoamyl: (25/24/1) alcohol extracted and the aqueous layer was treated with RNAse AþT1 for 1 h at 37°C. The lysate was again phenol/chloroform/isoamyl alcohol extracted and the gDNA was precipitated with equal volume of isopropanol in presence of 0.3 M sodium acetate at -20°C overnight. The gDNA was finally centrifuged, washed with 70% ethanol, air dried and reconstituted in 10 mM Tris-HCl, pH 8.0. And then, gDNA was denatured with 2 N NaOH at 42°C for 15 min, treated with sodium bisulfite at 55°C for 16 h, and purified by EZ DNA Methylation-Gold KitTM. RRBS libraries were constructed by standard protocols of the NuGEN Ovation® Ultralow Methyl-Seq Library Systems.
GEO
GSM3723830