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. 2004 Jul 19;32(13):3799–3806. doi: 10.1093/nar/gkh708

Figure 5.

Figure 5

The ETS-transcription factor, Pointed, binds to an ETS-binding site located 79 bp within ZAM LTR. The DNA binding domain of Pointed fused to GST (GST–PntC) was used in these assays. (A) The GST–PntC protein specifically binds the first 190 bp of ZAM. Increasing amounts of GST-PntC used in this gel-shift assay are 44, 88 and 264 ng. In comparison, no shift is observed with 250 ng of GST protein alone. (B) Competition experiments: binding is tested with a 264 ng of unlabeled GST–PntC, 100-fold excess of ZAMets and 100-fold excess of Mut an oligonucleotide mutated for the EBS (see Materials and Methods). (C) Competition experiments: the GST–PntC specifically binds the ZAMets oligonucleotide (from nucleotides 72 to 86 on ZAM sequence) which contains the presumptive EBS. A 100-fold excess of unlabelled ZAMets competes with the labelled oligonucleotide for the binding. A 100-fold excess of unlabelled oligonucleotide Mut, mutated for the EBS, is unable to compete. The asterisk and the arrow indicate the free probe and the gift, respectively.