840:153g:Projects/project22/2012/10/25

Attempted PCR and Gel Electrophoresis Verification

• In previous PCR runs, we did not obtain visible bands. We suspect that this was because the annealing time was not long enough. On Tuesday, we began another run of PCR with a 3 minute long annealing time. It was later discovered that the PCR cycle order was incorrect, so we then attempted to run the PCR again with the correct cycle orders. Today, we ran a gel on our PCR products to see if the PCR worked or not.

Lane 3 marker on the right to Lane 12 marker on the left

As you can see in the above image, our PCR products (the lanes between the markers) did not show up (even the positive control). This is most likely due to our erroneous PCR cycling. We could confirm the negative results of today's electrophoresis by running another PCR and gel electrophoresis check, but we have had poor results (gel electrophoresis checks with questionable DNA content - I.E. results indistinguishable from random proteins) with this DNA since day 1, so we will start preparing for another DNA extraction next week.

• We also began collecting buffers and the necessary materials for another DNA extraction for next week, where we will use the OpenWetWare extraction protocol.