GSM4730177: HEK293T-TTseq-rep2; Homo sapiens; OTHER
Total RNA was extracted using Trizol. The CUT&Tag tagmentation products were purified with 1X Sera-Mag carboxylate-modified magnetic beads (Sigma-Aldrich, GE24152105050350), and were eluted in 10 μL 0.1% Tween-20. The eluent was mixed with 10 U Bst 2.0 WarmStart DNA polymerase (NEB, M0538) in 1X Q5 polymerase reaction buffer, and was incubated at 65 °C for 30 minutes to complete the strand displacement reaction. The reaction was then stopped by incubation at 80 °C for 20 minutes. To perform the library preparation, the mixture was mixed with a universal i5 primer and a uniquely barcoded i7 primer, and was amplified with Q5 high-fidelity master mix (NEB, M0492). The libraries were size-selected with 0.56-0.85´ Sera-Mag carboxylate-modified magnetic beads, and were subjected to LabChip DNA analysis and Illumina sequencing. The  RNA was purified with the Sera-Mag carboxylate-modified magnetic beads and was subjected to strand-specific RNA-seq library preparation. Libraries were made with the NEBNext ultra RNA library prep kit for Illumina and subjected to next-generation sequencing.