Figure 8: Timeline of experimental protocols for gene knockdown, showing larval development, dsRNA expression, and wounding.

A) For all basement membrane source experiments, animals were maintained at 25°C. Larvae developed to early 3^rd instars, were pinched, and usually recovered 24 h before dissecting. For experiments where no wound was inflicted, larvae developed to late 3^rd instar prior to dissecting. B) For basement membrane hierarchy of repair and function experiments in which LanB1, vkg, or trol was knocked down, embryos were laid and allowed to develop to 1^st instar larvae at 18°C, with dsRNA not expressed. During 1^st instar, bottles were shifted to 29°C to promote dsRNA expression, and larvae developed to early 3^rd instar prior to wounding. After wounding, larvae recovered for 24 h at 29°C prior to dissecting. For Ndg KD experiments, bottles were maintained at 29°C for the entire experiment (not shown). Only control, trol KD and Ndg KD larvae were capable of pupariating. dsRNA^BM denotes dsRNA against vkg, trol, or LanB1.