Team:Warsaw/Calendar-Main/10 September 2009

Assembly of fusion proteins Marcin Task 1: Methods: 3 &mu;l purified plasmid DNA product 0.5 &mu;l XbaI (Fermentas) 0.5 &mu;l PstI (Fermentas) 2 &mu;l Buffer Tango (Fermentas) 14 &mu;l MQ water Results:
 * Digest plasmids isolated in 09.09.2009 to reveal the identity of the constructs
 * Reaction mixture composition:
 * Digest was performed 90 minutes
 * None constructs are correct. It is obligated to perform cloning second time. If I dephosphorylate the vector prior to ligation the result should be better.

Task 2: 15 &mu;l purified plasmid DNA product 2 &mu;l Tango Buffer (Fermentas) 1 &mu;l CIAP (Fermentas) Cloning of the mgtc promoter into the pSB1A3 plasmid Kamil
 * dephosphorylation the following molecules previously digested by resctriction enzymes:
 * BBa_K177036
 * pKSII cloning vector
 * Reaction mixture composition:
 * Reaction was performed 45 minutes and subsequently inactivate via heating (65 &deg;C)

Tasks:  Plasmid isolation Control digest 

Methods:  Plasmids from 3 ml of overnight culture were isolated using the Plasmid Mini kit (A&A Biotechnology). The digest mix was prepared as follows: 20&mu;l isolated plasmid 3&mu;l Buffer O (Fermentas) 1&mu;l PstI enzyme 1&mu;l EcoRI enzyme 5&mu;l H2O  The digest was carried out for 3h in 37&deg;C and then inactivated in 80&deg;C for 15min. The results were visualised on 1% agarose gel 

Results:  <img src="http://2009.igem.org/wiki/images/c/c5/2009.09.10_-_mgtC.jpg"> All empty</li> </ul> Conclusions:  So tired, need sleep... </li> </ul>

Cloning of the cro-box into the pSB1A3 plasmid Kamil

Tasks:  Plasmid isolation</li> Control digest</li> </ul>

Methods:  Plasmids from 3 ml of overnight culture were isolated using the Plasmid Mini kit (A&A Biotechnology).</li> The digest mix was prepared as follows: 20&mu;l isolated plasmid 3&mu;l Buffer O (Fermentas) 1&mu;l PstI enzyme 1&mu;l EcoRI enzyme 5&mu;l H2O </li> The digest was carried out for 3h in 37&deg;C and then inactivated in 80&deg;C for 15min.</li> The results were visualised on 2% agarose gel</li> </ul>

Results:  <img src="http://2009.igem.org/wiki/images/a/a9/2009.09.10_-_cro-box.jpg"> All empty</li> </ul> Conclusions:  So tired, need sleep... </li> </ul> I actually wrote some very witty comments bellow every picture, but it seams that two people can't edit one page at the same time and I got an error when Marcin saved his. Believe me, the comments were hilarious, but now I'm just to tired to write them again so you have to be satisfied with the "all empty" notes.