α h3k9me3

Imaginal discs were dissected and fixed in 4% paraformaldehyde for 20 min, washed and permeabilized in phosphate-buffered saline with 0.1% Triton X-100, and blocked in 10% normal goat serum. Primary antibodies used were α-Ci [1:25; Developmental Studies Hybridoma Bank (DSHB)], α-Ubx (1:10; DSHB), α-En (1:25; DSHB), α-β-galactosidase (1:500; Promega), α-V5 (1:500; Sigma-Aldrich), α-Wg (1:100; DSHB), α-Cut (1:100), α-Ptc (1:50; DSHB), α-pMAD (1:500; Abcam), α-Smo (1:10; DSHB), α-MMP1 (1:100; a combination of 14A3D2, 3A6B4, and 5H7B11; DSHB), α-Tara (1:700; K. Koh), α-H3K27me3 (1:500; Active Motif), α-H3K9me3 (1:500; Active Motif), α-H3K4me1 (1:500; Active Motif), α-H3K4me3 (1:500; Active Motif), α-H4ac (1:500; Active Motif), and α-Sd (1:500; K. Guss). Secondary antibodies were from Cell Signaling Technology and used at 1:400. Nuclei were stained with 4′,6-diamidino-2-phenylindole (1:1000; Cell Signaling Technology). Samples were imaged on a Zeiss LSM 700 confocal microscope.