Fig. 2.

Characterization of antimicrobial peptide induction following pinching larvae with forceps. (A-C) Real-time qPCR analysis of Drs expression (A,C) or Dpt expression (B) upon clean injury or pinching larvae with forceps at the indicated time points with larvae of Oregon R, w¹¹¹⁸ and y w. Pinching was performed with larvae of w¹¹¹⁸ in C. (D) Melanization spots after pinching larvae with forceps after 4 h. The indicated magnification of the objective lens was used; arrowheads indicate melanization spots. The right bar graph shows the percentage of larvae that exhibited melanization spots. Data were analyzed by Student's t-test and values represent the means±s.e. of three independent experiments with 30 larvae each. (E) Drs-GFP reporter analysis using a fluorescence stereomicroscope upon pinching larvae with forceps after 4 h. Larvae of Oregon R, Drs-GFP Dpt-lacZ (DD1); c564-GAL4, DD1; Dif¹ and DD1; Dif^nmc were used. Brightfield (top row) and fluorescence images (bottom 2 rows) of single larva or multiple larvae (bottom row); the green signal indicates GFP fluorescence. (F) Real-time qPCR analysis of Drs expression following pinching of Oregon R, w¹¹¹⁸, and y w larvae with forceps at the indicated time points. The larval fat body was dissected out from other tissues. (G,H) Real-time qPCR analysis of bacterial genomic DNA coding for 16S rRNA, which was normalized by the Drosophila genomic region for 12S RNA from conventionally reared or germ-free Oregon R larvae before pinching (G), or of Drs expression following pinching larvae with forceps at 4 h with conventionally reared or germ-free Oregon R larvae (H). Data were analyzed by the Student's t-test and values represent the means±s.e. of three independent experiments with 10 larvae each (F,G). *P<0.05.