Generating asp null allele using CRISPR. (A) Schematic of the asp locus and region targeted for deletion. Position of guide RNAs (red arrows) and primers used for PCR screening (orange arrows; S.P., sequence primers). TSS, transcriptional start site. (B) PCR screen from control (yw) and asp^t25/Df flies. (C) Quantitative PCR of asp transcript levels (three biological replicates, error bars are SEM). (D) Head size in age-matched control (TM6B) and asp^t25/Df adults. (E) NBs from control (top panel, asp^t25/TM6B) and asp^t25/Df (bottom panel) mutant larvae expressing GFP-CaM and stained for β-tubulin, pH3, and centrosomin (Cnn). White arrowheads denote pole position and red arrowheads denote centrosome position. Bars: (D) 1 mm; (E) 2 µm.