840:153g:Projects/project23/2012/10/18

PCR Amplification of g. sulfereducens DNA

• On Tuesday, we amplified or gene of interest from geobacter sulfereducens DNA. Since we need to remove an internal restriction site in our gene, we preformed a point mutation. This was done by amplifying two reactions. This creates two sections with an small overlapping sequence over the mutation site. Our oligos contain a base-change so that an overwhelming majority of the PCR product should not have the restriction site any longer. We also ran each reaction at three different annealing temperatures since our particular oligos are potentially problematic.
• On Thursday, we ran our reactions and controls from Tuesday on a 1% agarose gel to check for intended PCR product.