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Determination of free and core-bound σ32 by coimmunoprecipitation. Core RNAP was captured by bead-immobilized anti-β and -β′ antibodies. SDS-PAGE gels were run on fractions containing free (A) and bead-immobilized (B) σ32 (indicated by arrows). The identities of the σ32 are shown above each lane. After transferring the proteins to nitrocellulose membranes, these were probed with polyclonal anti-σ32 antibodies to reveal the presence of σ32. See Materials and Methods for details.
