Self-doubt kills talent

Self doubt kills talent and yesterday...it almost killed my experiment!
I'm not too sure why it happens but I'm getting better at recognizing it!

This week, I was able to get my own personal DNA extracted from buccal cells and started a very rudimentary version of my DNA extraction experiment.  I wanted to be sure that I could pull it off before I started with the really expensive materials that were ordered for my experiment.  It took me 3 tries to get my gel for the electrophoresis set up correctly, but you know what they say, the third time is the charm! 

I followed an old protocol that I had from last semester for running the gel.  I made my control dye which consisted of 9 um (microliters) of molecular mass ruler and 1 um of SYBR green nucleic acid gel stain and added it to my first well of the gel.  Then, with three different buccal samples, I added 9 um of the extracted DNA, 1 um of SYBR green and 2 um of orange loading dye. I placed them in wells 3, 5 and 7 and filled the electrophoresis rig with 1x TAE buffer solution, hooked it up to the voltage machine and ran 120 volts through it for 40 minutes.

This is what my gel looked like when it was finished and it is also where my doubt set in!

(the blue color is my control and the 3 faint orange colors are from the orange loading dye)

Maybe I have been watching too many episodes of CSI but some some reason I was expecting to see dark bands of DNA that ran down my gel.  I felt defeated to say the least and I was about to toss out my gel when Josh told me to have Matt take a look at it.  It was perfect timing as Matt and Anil were talking and they came over to see my progress and boy....does it pays to ask the experts!  It worked!!  The bands that I was expecting to see would not show up until I ran my actual PCR, (phew).  Anil put my gel in the Bioimaging system which shines a UV light onto the gel in order for the maker, DNA and loading dyes to glow.

This is what was produced:

 (On the left side of the gel you can see 3 bright bands towards the top, that is my DNA, they are glowing).

From here, my next step will be to run the PCR and amplify (or copy) the DNA sequence from those samples.  It is from that process, that I will be able to see the rows of dark bands form on a similar gel.