Team:Warsaw/Calendar-Main/15 April 2009

Cloning of the mgtc promoter into the pKSII+ plasmid Kamil

Tasks:  Amplification of mgtc and hly 

Methods:   PCR mixture's composition: 2ul buffer 1ul MgCl2 0,5ul primers 1,5ul dNTPs (10 mM) 01,ul polymerase Solution was topped up with H2O to 20ul. 2 repeats of every sample were made.    PCR programs: mgtc 90s 95&deg;C (30s 95&deg;C, 35s 48&deg;C, 60s 72&deg;C)x2 (30s 95&deg;C, 35s 58&deg;C, 60s 72&deg;C)x28 600s 72&deg;C ~ 4&deg;C hly 90s 95&deg;C (30s 95&deg;C, 35s 42&deg;C, 150s 72&deg;C)x2 (30s 95&deg;C, 35s 47&deg;C, 150s 72&deg;C)x28 600s 72&deg;C ~ 10&deg;C   Electrophoretic separation on 1% agarose gel 

Results: <img src="http://2009.igem.org/wiki/images/7/70/2009.04.15_-_PCR_mgtc_i_hly_opisane.jpg"/>  Gel (from left)</li> </ul> <ol> GeneRuler DNA Ladder Mix #SM0333 (Fermentas)</li> mgtc 1 repeat</li> mgtc 2 repeat</li> mgtc control -</li> hly 1 repeat</li> hly 2 repeat</li> hly control -</li>

</ol>

Notes:  Inappropriate template DNA was used for mgtc (ganomic DNA from Yersinia instead of genomic DNA from Listeria).</li> </ul>