Stability of lac represser-operator complexes in a new agarose-based gel matrix

摘要

Interactions of proteins with nucleic acids are central to many cellular processes including DNA replication, recombination and repair, and transcription. A powerful technique for the study of protein-nucleic acid interactions is the gel mobility-shift assay (1,2). In this assay, protein-DNA complexes are separated from the free nucleic acid and protein components by non-denaturing gel electrophoresis. The potential to resolve complexes differing in stoichiometry (1,3) and/or conformation (4-6) are major advantages of this procedure. The most widely used gel matrix for mobility shift analyses is polyacrylamide. Polyacrylamide gels are capable of excellent resolution of small complexes (M_r < 0.5 ×10~6) and may stabilize protein-DNA complexes duringelectrophoresis (1,7,8). Agarose gels are useful for studies of larger species (c.f., 9-12), although some complexes are significantly less stable in agarose than in polyacrylamide gels (13). This report describes a new agarose-based gel matrix, MetaPhor XR, that stabilizes lac represser-operator complexes to an extent similar to that found in 10% polyacrylamide gels.