Team:Warsaw/Calendar-Main/29 August 2008

Purification of proteins: A-alpha

Piotr Transformation of Rosetta with pET15b+A-alpha plasmid. Plating on LB with chloramphenicol and ampicillin.

Blue-white screening and rifampicin test in GM2163 Michał L.

GM2163 competent cells carrying pZC320 were transformed with following plasmids:

pMPMT5-omega, pMPMT5-AID,</li> pMPMT5-AIDT7</a>, </li> pMPMT5-AID+T7</a>.</li></ul>

Transformants were selected on CTAXI plates (LB + Chloramphenicol + Tetracycline + IPTG + Ampicillin 30 μg/mL + X-Gal) because GM2163 are chloramphenicol resistant.

Cloning of protein A DNA to pET15b+OmpA-alpha</a> plasmid Michał K.  <ol>Isolation</a> of pET15b+A-alpha</a> plasmids.</li>  Control digest</a> of isolated plasmids with XbaI and BamHI (Tango buffer). </li> Gel electrophoresis (Fig. 1.</a>). Choice of proper clone. </li> </ol> <img src=" http://2008.igem.org/wiki/images/a/a8/Kokodak.jpg" width=300/></a> Fig. 1. Control XbaI/BamHI digests of plasmids isolated from transformants with colony PCR product. 1. Marker 2-4. Digested plasmids pET15b+A-alpha