GSM5426085: BJAB-SCNG-H3K27ac-Rep1; Homo sapiens; OTHER
BJAB cells CUT&Tag was carried out following manufacture's instruction (Vazyme Biotech, TD901-01). In brief, transient transfection of EBNA2 and vehicle plasmid into BJAB cell respectively, collect positive cells by fluorescence-activated cell sorting (FACS) after 48 hours. Approximately 100,000 BJAB cells were processed by centrifugation between buffer exchanges at 600×g for 3 min and in a swinging bucked rotor at room temperature for the initial wash and incubation steps. Cells were collected and washed with 1 ml of Wash Buffer at room temperature. Cells were rotate and incubated in antibody (ab4729, Abcam, USA) diluted 1:50 in Antibody Buffer for 2 hours at room temperature. Permeabilized nuclei were then centrifuged once and incubated with anti-rabbit IgG antibody (FD0218, Fudebio, China, 1:100 dilution) in 1 ml of Dig-wash Buffer on a rotator at room temperature for 1 hour. Nuclei were washed twice with Dig-wash Buffer and incubated with 1:100 dilution of pG-Tn5 in Dig-300 Buffer for 1 h at room temperature on a rotator. Cells were washed two times with Dig-300 Buffer and resuspended in 300µl of Tagmentation Buffer and incubated at 37 °C for 1 h. Tagmentation was stopped by adding Stop Buffer (each sample with 10μl 0.5M EDTA, 3μl 10% SDS and 2.5μl 20mg/ml Proteinase K), and the sample was held for 1 hour at 50 °C in incubator. The remaining steps are carried out according to the product manual.