Slowly but surely

I ended my project last week with great extractions, and with the conclusion that a 24 hour culture is the ideal time I need for bacteria to grow in, so I can extract DNA from them. What I began to do on Tuesday was my PCR,  even though last time my PCR results showed that primer 7 did not work at all and primer 3,4 and 6 only worked for a few bacteria I decided to work with these four primers once again. The bacteria I used were the six listed below. My results were awesome for primer 6! Those are the results I have been aiming for this whole time. I noticed that primer 3 kind of work but because the gel I ran it in seemed a little weird I cannot really conclude anything from this. Primer 4 and 7 worked for only 2 bacteria, each different from the other. I think the reason my PCR results were better this time around is because for one the practice; the more I do it the better they should be, and two I made sure all the reagents for this protocol were on ice.

DNA

• E. coli
• B. subtilis
• S. aureus
• Salmonella
• S. marcescens
• P. stuartii  

PCR Protocol

1. 10 microliters of master mix
2. 5 microliters of DI H2O
3. 4 microliters of primer (3,4,5,6)
4. 1 microliter of DNA 

Primer 3

Primer 4

Primer 6

Primer 7

RFLP Results using DNA extractions from 4-11-13

       RFLP Protocol

      1. 10 microliters of 1 x buffer

      2. 88 microliters of DNA

      3. 2 microliters of Ecor 1 enzyme

For the RFLP I used the same DNA extractions that were done on 4-11-13, and the same protocol I used for the RFLP done on 4-4-13. My results were what we would call sloppy, because you can see the DNA and you can see where the markers are but the DNA is just smeared all down the lane. We are now trying to find a way to lower the concentration of DNA because we think the reason we keep getting bright lanes all the way down is because the DNA concentrations are high.

I also want to add that on Friday I was able to experience the wonderful Desert Botanical Garden, it was awesome and I got to learn the plants we have here in Arizona. Thanks to my advisors, Dijana, and Schampel I was able to learn so much about these plants and reinforce some information I was introduced to in my biology classes. I also want to share with you the plant the cactus that made my day at this garden. This cactus is called the old man cactus.