Quantstudio 7 flex real real time pcr system

Total RNA of collected tissues were extracted with TRIzol RNA Isolation Reagents (Thermo-Invitrogen) and reverse-transcribed with HiScript II Q RT SuperMix for qPCR (Vazyme). Real-time PCR was performed with FastStart Universal SYBR Green Master (Roche) and with different primer sets on QuantStudio 7 Flex Real Real-Time PCR System (Applied Biosystems). The primers used for real-time PCR are shown in Supplementary Data 1. The 2–ΔΔCt method was used to calculate relative expression changes.

The ChIP assays were performed using ChIP kit (Catalog no. 17-371; Millipore). The procedure was according to the kit instruction manual provided by the manufacturer. Briefly, 1 × 10⁷ Myc-CaP cells were fixed by 1% formaldehyde, fragmented by sonication to shear the chromatin to 400–1000 bp. The sheared crosslinked chromatin was incubated with IgG, anti-BRG1 (Abcam, clone: EPNCIR111A, ab110641), anti-BAF155 (Santa Cruz Biotechnology; clone: G-7, sc-365543X), anti-ARID1A (Cell Signaling Technology; clone: D2A8U, 12354 S) and anti-ARID1B (Cell Signaling Technology; clone: E9J4T, 92964 S) antibodies (10 μg antibody for each ChIP reaction) overnight followed by Protein G conjugated agarose beads incubation. The precipitated DNA was amplified by primers and quantified by QuantStudio 7 Flex Real Real-Time PCR System (Applied Biosystems). ChIP primer sequences can be found in the Supplementary Table 3.

Total RNA was extracted using TRIzol (Invitrogen) according to the manufacturer’s instructions. First-strand cDNA was synthesized by HiScript II Q RT SuperMix (Vazyme) for qPCR. Real-time PCR was performed with FastStart Universal SYBR Green Master (Roche) on QuantStudio 7 Flex Real Real-Time PCR System (Applied Biosystems). The primers used for real-time PCR are shown Supplementary Table 3. The 2–ΔΔCt method was used to calculate relative expression changes.