Cleavage under targets and tagmentation

The CUT&Tag protocol was described previously (43). The Hyperactive In-Situ ChIP Library Prep Kit was purchased from Vazyme, and libraries were generated following the manufacturer’s recommendations. Briefly, 1 × 10⁵ cDC1s or cDC2s sorted from the spleen of WT or Zfp366^−/− full chimeric mouse were bound to concanavalin A–coated magnetic beads (Bags Laboratories) and were subjected for immunoprecipitation with 0.5 μg of primary antibody [anti–DC-SCRIPT or rabbit anti-mouse IgG (immunoglobulin G) control]. After primary incubation and washing using a magnetic stand, a secondary anti-rabbit antibody was added and incubated under gentle agitation for 1 hour (see table S6 for antibody details). Cells were washed and incubated for 1 hour in a mix of hyperactive pG-Tn5/pA-Tn5 transposon with Dig-300 buffer to a final concentration of 0.04 μM. Cells were washed and resuspended in 100 μl of tagmentation buffer (10 mM MgCl₂ in Dig-300 buffer) and incubated at 37°C for 1 hour. Reaction was stopped by heat inactivation (55°C for 10 min) and DNA purified using AMPure XP beads (Beckman Coulter). For library amplification, 24 μl of DNA was mixed with 10 μl of 5× TruePrep Amplify Buffer (TAB), 1 μl of tris-acetate-EDTA, and 5 μl of uniquely barcoded i5 and i7 primers (67) and amplified for 14 cycles. PCR products were purified with AMPure XP beads and eluted in water. Libraries were sequenced on an Illumina NextSeq platform, and 150-bp paired-end reads were generated.