Team:Warsaw/Calendar-Main/18 August 2009

Assembly of endosomal detection operon Marcin Task 1:  Gel-outs of following constructs digested on 17.08.09: p53 CDS on  pSB1A3 plasmid  BBa_R0080 on  pSB1A3 plasmid  BBa_E0010</a> on  pSB1A3</a> plasmid</li>  BBa_C0040</a> with  RBS</a> on  pSB1A3</a> plasmid</li></ul></ul> Methods: <ul> All samples were inactivated via heating in 80 &deg;C for 20 minutes All gel-outs were performed using the EurX gel-out kit according to the manual</a></li></ul> Results: <img src="http://2009.igem.org/wiki/images/7/7d/PSB_p53_R0080_C0040%2BRBS_gel_out_ocena_18_08_09.png"> <ul>All gel-outs (except C0040 with RBS) were successful. One isolation failed because of very low DNA concentration</ul></li> Task 2: <ul> Digest pKSII plasmid with cro CDS</li></ul> Methods: <ul>  Reaction mixture composition:</li> 45 &mu;l purified plasmid DNA product 1 &mu;l XbaI (Fermentas) 0.5 &mu;l SpeI (Fermentas) 5 &mu;l Buffer Tango (Fermentas) </ul> Task 3: <ul> Ligate p53 CDS to  pSB1A3</a> plasmid with  BBa_B0032</a> </li></ul> Methods: <ul> Reaction mixture composition:</li> 10 &mu;l p53 CDS 7 &mu;l vector 2.3 &mu;l Buffer Tango (Fermentas) 3 &mu;l dNTPs mixture (EurX) 1 &mu;l T4 ligase (Fermentas) Ligation was performed for about 9 hours and it was subsequently thermally inactivated</li> </ul>

Making of plac-RBS-llo part Jarek

Tasks: <ul> Preparation of 20 liquid cultures, from colonies that grew from ligation, in 3 ml of LB+Amp nad incubating them in 37C degree</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: <ul> <li> </li> </ul>