Flag-CUT_RUN in wild-type and 3xFlag-Morc2 germ cells

To investigate the genomic targets of Morc2 in different germ cell types, Flag-CUT_RUN was performed on pachytene spermatocytes (Pac) from 3xFlag-Morc2 mice, P12 testicular cells from 3xFlag-Morc2 mice, and their corresponding wild-type controls. Experiments were conducted in biological triplicates following the manufacturer protocol for the CUTANA ChIC_CUT_RUN Kit (Cat. No. 14-1048-24, EpiCypher). Briefly, 500,000 cells were collected and bound to Concanavalin A-coated beads for 10 min at room temperature. The bead-cell complexes were incubated with 1 ug anti-FLAG primary antibody in 0.01% digitonin buffer. After washing, Ca2+-activated pAG-MNase was added to cleave DNA at antibody-bound sites. CUT_RUN DNA was extracted and purified using SPRIselect beads, followed by end repair, adaptor ligation, U excision, and PCR amplification using the CUTANA CUT_RUN Library Prep Kit (Cat. No. 14-1001, EpiCypher). Libraries were sequenced on an Illumina NovaSeq 6000 platform to generate paired-end 150 bp reads at Azenta Life Sciences.