Team:Warsaw/Calendar-Main/23 August 2009

Assembly of endosomal detection operon Marcin

Task 1:  Transformation of chemocompetent E. coli strain DH5&alpha Constructs to transform:  Bax CDS +  BBa_B0032  phoP/Q CDS +  pSB1A3 plasmid Methods:  Detailed protocol of transformation is described <a href="http://2009.igem.org/Team:Warsaw/Calendar-Main/5_July_2009">here</a>.</li></ul> Task 2:  Isolate of plasmids from bacterial cultures prepared in <a href="http://2009.igem.org/Team:Warsaw/Calendar-Main/22_August_2009">22.08.09</a></li></ul> Methods:  Plasmids were isolated using the A&A plasmid mini kit. Detailed procedure of the isolation is described <a href="http://aabiot.com/products/dna_purification/plasmid_dna/plasmid_mini/protocol_plasmid_mini.pdf" here>here</a></li></ul> Task 3:  Digest previously isolated plasmids and verify the correctness of the ligation:</li></ul> Methods:   Reaction mixture composition:</li> 2 &mu;l purified plasmid DNA product 0.5 &mu;l XbaI (Fermentas) 0.5 &mu;l PstI (Fermentas) 2 &mu;l Buffer Tango (Fermentas) 15 &mu;l MQ water Digest was performed one hour</li> Results: <img src="http://2009.igem.org/wiki/images/5/56/R0080_C0040%2BRBS_23_08_09.png" width="45%" heigth="45%"> <font face="Times New Roman" size="3"> All isolated samples have expected plasmid

Making of the plac-RBS-llo-intA part Jarek

Tasks:  Transformating the competent E.coli strain TopF' with ligation</li> Plating the transformated culure on LB+Amp plates</li> </ul>

Cloning switch 1 regulatory parts [ <a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K177012">K177012</a>-PcI.RBS.LacI, <a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K177033">K177033</a>-PcI.RBS.LacI.PcI.RBS.RFP.terminator, <a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K177011">K177011</a>-PLacI.RBS.cI.terminator, <a href="http://partsregistry.org/Part:BBa_K177038">K177038</a>-PLacI.RBS.cI.terminator.PLacI.RBS.GFP.terminator ] into two compatible low copy number plasmids of different antibiotic resistance Ania Tasks:   </li> </ul>