10.1371/journal.pone.0006652.g003 Neha Agrawal Neha Agrawal Nisha Padmanabhan Nisha Padmanabhan Gaiti Hasan Gaiti Hasan Delayed pupation in <i>Dilp2GAL4</i> and <i>DdcGAL4</i> rescue conditions results in larger body size. Public Library of Science 2013 pupation conditions larger 2013-02-21 05:02:20 Figure https://plos.figshare.com/articles/figure/_Delayed_pupation_in_Dilp2GAL4_and_DdcGAL4_rescue_conditions_results_in_larger_body_size_/558537 <p>(A) From left to right: <i>UASitpr<sup>+</sup>/+;Dilp2GAL4/+; P0163GAL4,itpr<sup>sv35/ug3</sup></i> (<i>Dilp2GAL4 P0163GAL4</i> rescue), wild-type (<i>Canton-S</i>) and <i>UASitpr<sup>+</sup>/+</i>;<i>Dilp2GAL4/+</i>;<i>itpr<sup>sv35/ug3</sup></i> (<i>Dilp2GAL4</i> rescue) pupae. (E) Only <i>UASitpr<sup>+</sup></i> expression with <i>Dilp2GAL4</i> in <i>itpr<sup>sv35/ug3</sup></i> background causes a significant increase (*p<0.005; Student's t-test) in pupal length as compared to controls of all the indicated genotypes. Pupal length is restored close to wild-type in <i>Dilp2GAL4::P0163GAL4</i> rescue condition. (B) From left to right: wild-type and <i>Dilp2GAL4</i> rescued flies. (F) Body length of <i>Dilp2GAL4</i> rescued flies is significantly more than wild-type flies (*p<0.05; Student's t-test). (C, D) From left to right: <i>UASitpr<sup>+</sup>/+;DdcGAL4/+;P0163GAL4,itpr<sup>sv35/ug3</sup></i> (<i>DdcGAL4 P0163GAL4</i> rescue), wild-type (<i>Canton-S</i>) and <i>UASitpr<sup>+</sup>/+</i>;<i>DdcGAL4/+</i>;<i>itpr<sup>sv35/ug3</sup></i> (<i>DdcGAL4</i> rescue) pupae (C) and flies (D). (G) Only <i>UASitpr<sup>+</sup></i> expression with <i>DdcGAL4</i> in <i>itpr<sup>sv35/ug3</sup></i> background causes a significant increase (*p<0.005; Student's t-test) in pupal length as compared to controls of all the indicated genotypes. Pupal length (G) and adult weight per fly (H) is increased in the <i>DdcGAL4</i> rescued condition (*p<0.005; Student's t-test) but is restored close to wild-type in <i>DdcGAL4::P0163GAL4</i> rescue condition. n = 10 or more for each individual genotype for (A–G). For (H), the following numbers of male flies (n) in batches of around 3 flies each were weighed for each genotype: <i>DdcGAL4 P0163GAL4</i> rescue n = 26, wild-type n = 30 and <i>DdcGAL4</i> rescue n = 54. (I) Total number of larvae that undergo pupation is significantly increased on introducing a prothoracic gland GAL4 (<i>P0163GAL4)</i> in <i>DdcGAL4</i> and <i>Dilp2GAL4</i> rescued conditions (*p<0.05, **p<0.005; Student's t-test). (J) Time AEL for 50% pupal formation is significantly reduced with <i>P0163GAL4</i> in <i>DdcGAL4</i> and <i>Dilp2GAL4</i> rescued conditions (*p<0.005; Student's t-test). However the 50% pupation time in all single and double GAL4 conditions remained longer than the 50% pupation time of wild-type. For (I) and (J) 25 larvae in the following number of batches (N) were assayed for pupation for each genotype: wild-type N = 11, <i>Dilp2GAL4</i> rescue N = 10, <i>Dilp2GAL4 P0163GAL4</i> rescue N = 5, <i>DdcGAL4</i> rescue N = 13 and <i>DdcGAL4 P0163GAL4</i> rescue N = 9. (K) <i>DdcGAL4</i> rescued <i>itpr<sup>sv35/ug3</sup></i> larvae pupated earlier on being fed 20-hydroxyecdysone (∼150 hrs AEL) than larvae without 20-hydroxyecdysone (∼200 hrs AEL) (*p<0.05; Student's t-test). A minimum of 75 animals were screened in batches of 25 each. Differences in pupation rate are not apparent upon 20-hydroxyecdysone feeding in <i>Dilp2GAL4</i> rescue animals due to increased lethality in this condition in late third instar larvae. The <i>DdcGAL4</i> rescued condition which were not fed ecdysone, pupated at a slower rate than those observed in (J). This is very likely due to differences in culture conditions in the two cases. Results are expressed as mean±SEM.</p>