I am working on PIERCE lightshift chemiluminescent EMSA Kit.
As the attachment pic.
I use Promega TNTŪ Quick Coupled Transcription/Translation Systems to get my Protein truncation "A" from the literature sequence.(Positative control Luciferase translation was so good) .But when I put 2ul of my translated product together with NE into EMSA systerm, finally the membrane looks so dirty on the top in Lane 3. Lane 1 with NE only is ok.I think it is not the Pr "A" shift because Lane 2 was Luciferase product control with the same pattern.
How to avoid the contaminate?Should I purify the translated product?But Promega Kit said the product can be used dirrectly in EMSA assay.Or I put too much product in EMSA assay? Has anyone has the same problem?I will be so appreciated if anyone could give me some suggestion.