Journal: Scientific Reports

Article Title: Characterization of the MCM homohexamer from the thermoacidophilic euryarchaeon Picrophilus torridus

doi: 10.1038/srep09057

Figure Lengend Snippet: (A & B). Immunoprecipitation analysis. (A): PtMCM immunoprecipitates were resolved by SDS-PAGE (10%) and analyzed by western blotting using anti-MCM (to detect immunoprecipitated MCM) or anti-GINS (to detect co-immunoprecipitating GINS) antibodies (1:1000 dilution). Full-length blots can be seen on-line in . (B): PtGINS immunoprecipitates were similarly analyzed. Full-length blots can be seen on-line in . Lanes 1: antibody control (bead-bound antibodies only and no lysate added); lanes 2: lysate incubated with beads only (with no antibody coupled to the beads); lanes 3: immunoprecipitation reaction; lanes 4: molecular weight marker; lanes 5: input lysate. (C). MCM pulldown experiment: Coomassie stain of SDS-PAGE analysis. Lane 1-input purified PtMCM; lane 2 – input purified PtGINS; lanes 3 to 8 – analysis of eluate fractions of pulldowns. Lanes 3 and 6 - recombinant MCM alone incubated with Strep -Tactin Superflow affinity beads; lanes 4 and 7- recombinant GINS alone incubated with Strep -Tactin Superflow affinity beads; lanes 5 and 8- recombinant MCM plus recombinant GINS incubated with Strep-Tactin Superflow affinity beads. Mr – molecular weight marker.

Article Snippet: MCM (and any GINS that was interacting with it) was pulled down from the reaction using Strep-Tactin Superflow affinity beads (IBA BioTAGnology), by incubating the interaction reaction mix with the beads for 15 min at 4°C.

Techniques: Immunoprecipitation, SDS Page, Western Blot, Control, Incubation, Molecular Weight, Marker, Staining, Purification, Recombinant