GSM9077212: VSMCs_CON H4K16LA rep2; Homo sapiens; OTHER
The frozen tissues were first washed with pre-cooled 1X PBS, then thawed on ice, cut into pieces. Next, added appropriate amount of tissue lysate, and grinded them to homogenate. 2 ul from the homogenate was be used to observe the morphology and number of nuclei under microscope. After conforming the quality of nuclei, then strained the homogenate through 40 um/35 um cell sieve. Counted the nuclei from the filtered grinding solution. Finally, about 500,000 nuclei needed to be precipitated by centrifugation at 4 ℃ temperatures. After mixing the antibody I and Antibody Buffer in proportion, added it into the precipitate obtained in the above step, and incubated at room-temperature for 2h after mixing. The antibody II was added and incubated for 1 h at room-temperature. After centrifugation, the mixture of Dig Buffer and CUT&Tag-Tn5 enzyme was added and mixed, and incubated at room-temperature for 1 h.Purified the above products using DNA Purification Kit (MN，740609). To determine PCR cycle number, take a small amount of the purified product and do a pre_x0002_amplification.After the pre-amplified bands were correct, then done formal amplification. After determining the yield and the band type, purified and constructed the library.