Discs were dissected in PBS immediately upon downshift and collected as pools of 100 discs for early L3 samples and 50 discs for late L3 samples. The 4 samples were placed in lysis buffer (10mM Tris 7.5, 10mM NaCl, 3mM MgCl2, 0.1% IGEPAL CA-630), and pelleted for library prep. Each of the 4 samples was repeated in triplicate from separate repeats of larval ablation. ATAC-seq libraries were created using Illumina Tn5 transposase, essentially as in Buenrostro et al. 2013.