Hello everyone,

I would like to see the interaction between my protein of interest (tagging with GST) with DND probes, however I always get smears (as shown in the picture) after I run the gel. I am not sure what these are and how can I get rid of them, it would be great if I could get some suggestions.

What I did was I incubate my 1 uL of DNA probe (1500 nM) with 2 uL of *5x binding buffer and 7 uL of 60 ng of protein of interest. The reaction size is 10 uL. This binding reaction sits at RT for 15 mins. I then add 2uL of 150 mg Ficoll 400 before loading on the gel.I run the gel (3.5% acrylamide) in 0.5xTBE buffer for 40 mins in a cold room for 40-45 mins.

* 5x binding reaction contains 50 μL of 1 M Tris-HCl, pH 7.5; 10 μL of 5 M NaCl; 200 μL of 1 M KCl, 5 μL of 1 M MgCl2, 10 μL of 0.5 M EDTA, pH 8.0; 5 μL of 1 M DTT; 25 μL of 10 mg/mL BSA, and 695 μL of ddH2O. This is from a paper by Hsieh et al., 2016 (An Optimized Protocol for Electrophoretic Mobility Shift Assay Using Infrared

Fluorescent Dye-labeled Oligonucleotides)

Thank you so much!