Question

Mobility-shift assays can inform about protein-nucleic acid interactions by comparing:

a) the lengths of primer extension reactions in agarose gels.

b) elution times during affinity column chromatography.

c) sedimentation coefficients following gradient centrifugation.

d) the migration of molecular complexes in a non-denaturing polyacrylamide gels.

e) none of the above.

Answer 1

Ans is D- the migration of molecular complexes in a non denaturating polyacrylamide gels

In a mobility shift assay interaction of protein and nucleic acid is measured. In this assay using polyacrilamide gels usually three lanes are taken.

Lane 1 of the gel contains untreated genetic material (DNA or RNA)

Lane 2 contains the protein

Lane 3 contains the protein and the genetic material whose interaction needs to be checked.

Now when a protein will interact with a DNA it will produce a larger, heavier complex which will move slowly through the gel.

So if the DNA and protein molecule interact there will be a considerable shift in their band position from the other two lanes(Lane 1 and 2) after running the gel for a short period. This DNA protein complex will produce a lower band.

But if the interaction does not happen there can be seen two bands corresponding to DNA and protein individually.

For visualisation radioactive, biotin or fluorescent labelling is used.