840:153g:Projects/project27/2012/11/13

Failed PCR...Again

• PCR products from 11/8 were checked on gel electrophoresis. Again, no distinct bands were shown by the products amplified by either primer set. The negative control, in which water was used instead of primers, also showed no amplification. The positive control, in which a known template and primer set was used, showed a distinct band at ~500bp. The two controls indicate that the PCR procedure was adequate for amplification and that there was no significant contamination. Lanes that included a 1:10 dilution of template DNA showed no amplification. Lanes that included 5μL template DNA and lanes with products with annealing temperatures of 45°C, 47°C, and 50°C all showed smearing with no distinct bands visible.

Lanes
 1,6,13: Marker
      2: 5μL template, mmoXY primers, 60.7°C anneal
      3: 5μL template, mmoBZDC primers, 62°C anneal
      4: 1:10 template dilution, mmoXY primers, 60.7°C anneal
      5: 1:10 template dilution, mmoBZDC primers, 62°C anneal
    7-9: mmoXY primers, anneal of 45°C, 47°C, and 50°C, respectively
  10-12: mmoBZDC primers anneal of 45°C, 47°C, and 50°C, respectively
     14: negative control (no primers used)
     15: positive control (known template and primers)