Purification and Histone Methyltransferase Assay of HAP

Full length open reading frame of HAP was amplified from the pGWB402 vector using the primers listed in Supplementary Table S5 and cloned into the pGEX4T-3 expression vector. The resulting plasmid was transformed into E. coli BL21-CodonPlus-RIL cells (Agilent) and protein expression assays were then performed as previously described [58 (link)]. Briefly, cells containing pGEX4T-3 vector were grown to an exponential phase and protein expression was induced by adding IPTG. Bacteria were then disrupted by sonication and protein purification was performed using GST-sepharose (Pharmacia) according to the manufacturer’s instructions. Finally, Histone H3 (K4, K9 and K27) methyltransferase activity was determined using HMT activity quantification assay kits (Abcam) following the manufacturer’s instructions.