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. 2016 Aug 2;6(10):3139–3148. doi: 10.1534/g3.116.032300

Figure 3.

Figure 3

Increasing UAS enhancer strength at the attP2 locus in trans increases attP2 UAS-PrP expression. UAS-mCD8::GFP constructs with variable UAS number were expressed at attP2 in trans to UAS-PrP, with expression driven by Cha-GAL4. (A) Western blot of fly head homogenates. Experiment was performed in triplicate with a representative blot shown. (B) Densitometry of Western blot signals in (A) and associated replicates. PrP signals were normalized to the kinesin loading control and then to the hemizygous attP2 UAS-PrP line lacking UAS-mCD8::GFP in trans. (C) Semiquantitative RT-PCR performed on fly head mRNA. Prnp and RpL32 transcripts were coamplified in the same PCR reactions, and each lane represents a biological replicate. CNTL is of mRNA prepared from UAS-PrP flies that do not express GAL4. (D) Densitometry of RT-PCR signals in (C). For each sample, the ratio of Prnp to RpL32 mRNA levels was taken and then normalized to the average value for the attP2 UAS-PrP line lacking UAS-mCD8::GFP in trans. Error bars in (B) and (D) represent standard deviations, with n = 3 for each group; ns denotes not significant, * P < 0.05, *** P < 0.001, and **** P < 0.0001. Control sample, CNTL; GFP, green fluorescent protein; mRNA, messenger RNA; PrP, prion protein; RT-PCR, reverse transcription polymerase chain reaction.