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. 2012 Nov 8;7(11):e47366. doi: 10.1371/journal.pone.0047366

Figure 5. GBX2 directly targets multiple genes associated with Usher syndrome and inner ear development.

Figure 5

(A) Reverse transcription (RT)-PCR analysis of Gbx2 and identified targets, Pcdh15, Ush2a, and Notch2, in E13.5 wild-type mouse cochlear or vestibular inner ear tissues. Myo15 positive control expression is observed in cochlear and vestibular tissues. (B,C,D) Gel-shift analysis for identified GBX2 targets USH2A, PCDH15, and NOTCH2. A reduction in the mobility of [ÿ -32P] ATP labeled USH2A, PCDH15, and NOTCH2 100-mer probes is observed with the addition of GBX2 (black arrows), whereas no shift is observed with the addition of GBX2ΔHD (compare lane 2 to lane 4). A supershift is observed in lane 3 with the addition of anti-GBX2. Addition of identical USH2A, PCDH15, and NOTCH2 100-mer unlabeled specific competitor probes at 100x, 300x, and 500x molar concentrations in lanes 5–7. Addition of USH2A, PCDH15, and NOTCH2 45-mer unlabeled non-specific competitor probe, omitting the GBX2 DNA-binding sequence in lanes 8–10.