GSM7993596: HUCCT1-HP1α; Homo sapiens; OTHER
CUT & Tag assay was performed with the Hyperactive Universal CUT&Tag Assay Kit (Vazyme). 1×106 TFK-1 cells were harvested and their nuclei were extracted. The extracted nuclei were immobilized with activated concanavalin A‐coated magnetic beads and reacted with anti-Flag antibodies (1:50) in 100 μL DIG buffer overnight at 4℃. After being washed, the bound antibodies were reacted with secondary antibodies (1:50) for 1 h at room temperature and incubated with 100 μl Hyperactive pA/G-Transposon adapter complex (0.04 μM) for 1 h at room temperature. The precipitated immunocomplex was diluted in Trueprep Tagment Buffer L and after proteinase K digestion, the DNA fragments were extracted and stored at -80 ℃. A DNA library was constructed using the TruePrep Index Kit V2 (Vazyme). The DNA fragments were mixed with N5XX and N7XX and amplified by PCR at 72 ℃ for 3 min, 95 ℃ for 3 min, and subsequent 10 cycles of 98 ℃ for 10 sec and 60 ℃ for 5 sec, 72 ℃ for 1 min and hold at 4 ℃. The library was extracted by VAHTS DNA Clean Beads (Vazyme). Average insert size of the molecules sequenced in A2 sample is 499. The specific gene sequences with paired ends were analyzed with DNA sequencing in an Illumina HiSeq 2000 platform (Illumina).