GSM5769311: si-ABCA1-seRNA-RXRα; Homo sapiens; OTHER
Libraries for high-throughput sequence were amplified with HyperactiveTM In-Situ ChIP Library Prep Kit for Illumina (Vazyme Biotech, TD903) according to the protocol. A universal adapter i5 and a uniquely adapter i7 primer gets from TruePrep Index Kit V4 for Illumina (Vazyme Biotech, TD501) with different barcoded for each sample. Using thermocycler with a heated-lid to amplify libraries under suitable conditions. PCR production purify was performed with 1.2X volume of VAHTS DNA Clean Beads (vazyme, N411-01) relative to PCR products according to the manufacture's. Finally, libraries were eluted with 22ul sterilized ultrapure water and separated by magnetic stand THP-1 macrophages were counted and harvested at room temperate. And then, cells were re-suspended with 500ul wash buffer and centrifuged for 3min at 600×g. Subsequently, cells were aliquoted 100ul in prepared ConA beads and incubated on a rotating platform for 10min at room temperate. Then, indicated primary antibodies and second antibodied were combined with cells. Hyperactive Tn5 transposon was applied to accurately cleavage DNA sequences near the target proteins. 300ul tagmentation buffer was added into product and incubated in 37°C for 1h to fragment the DNA sequences. At last, DNA was extracted for follow-up experiments