Extractions !

This week started off in a productive and interesting way. The reason I say this is because on Monday each of us had the chance to apply to another conference which will be taking place at Estrella Mountain Community College. Since the deadline was Monday we all had to revise our abstracts to submit them and have the chance to participate in this conference.

I am excited because my project changed a little bit, I will not be focusing on the best way to extract DNA anymore because I already found out what the best way is which was using the Epicentre QuickExtract kit, so what my project will focus on now is on the amount of successful PCR and RFLP results I will get from these extractions. In addition to this I will be extracting more bacteria then what I was working with.

On Tuesday I extracted DNA from the seven cultures I made last week. I noticed that the S. epidermidis bacteria did not grow so what Matt advise me was to set it aside and create another culture for it because it was obvious I was not going to be able to extract anything from it so I only extracted six bacteria. After my extractions were done I ran them in a .5 % agarose gel, but did not have great results so then I ran a new electrophoresis gel with 1% agarose to see if my results would change. I saw better results but I still lacked about three bacteria. Also I noted that instead of using normal 1x sybr green dye for running these gels I used 10x, this would help me because it should show really bright bands because of the increase in concentration.

Matt and I had a discussion the reasons why I had not gotten successful results for all six bacteria; we looked over my abstract and talked about maybe doing one of the steps different. This step included setting my extractions aside for an entire hour, then placing them in an 80 degree water bath for 2 minutes, and then continuing with the gel. I made sure to use the 1x sybr green dye that I usually use, and noticed how very light the color of the bands seemed compared to the first two results.

Figure 1 and 2 show the difference the concentration of agarose makes ( you can see the DNA better in Figure 2). Figure 3 shows the DNA in 1% agarose gel, but shows only a few bacteria.
   

Figure 1.
.5% Agarose Gel
Ruler
E. coli
S. aureus
B. subtilis
S. enterica
P. stuartii
S. marcescens
 

Figure 2
1% Agarose Gel
Ruler
E. coli
S. aureus
B. subtilis
S. enterica
P. stuartii
S. marcescens






Figure 3
1% Agarose Gel
Ruler
E. coli
S. aureus
B. subtilis
S. enterica
S. marcescens
P. stuartii

Back at it! :)

After being away from the lab for so long I glad to be back! I came in Monday to talk to my advisors about some internship opportunities over the summer. They were able to advise me about the qualities that someone looks for in an intern and gave some tips which cleared some questions up for me. Besides their awesome advice I was able to get some cultures done so when I would come in Tuesday I would have bacteria to work with. Since I am continuing with my  project my duty now is to test every bacteria I have access to in the lab. This means I will be working with new bacteria however I will continue to use the four main ones I have been working on. I prepared seven bacteria and incubated them over night these seven are:

• Salmonella
• S. Epidermidis
• Serratia Marcescens
• Providencia Stuartii
• E. coli
• Bacillus subtilis
• S. aureus

On Tuesday I was ready to try the Sigma and Epicentre protocol on these seven bacteria however I learned that we ran out of the reagents for each of the kits. I will have to wait until we order and receive them. Instead I tried the Isolation and Purification of Total Genomic DNA protocol and if you all recall this protocol was the least successful for my extractions. I tried it with only E. coli because it is always good to start fresh in an experiment and have an idea of what the outcome should be.  When I tried it back in December I only got DNA for E. coli, the other three bacteria were not visible after the electrophoresis gel run. I would like to try this protocol on these bacteria once more since I have been able to gain more experience in the lab and my skills have grown. Maybe this time I would be able to get positive results for DNA extraction.

 Also Josh gave me a new task to do after I do my DNA extractions instead of running a PCR I would do a Restriction Endonuclease Digestion on the DNA. This procedure is used to somehow cut this DNA sequence and using PCR we can reproduce some of the chunks of sequence that would be missing. I still do not have a clear understanding of this new part of my experiment but my homework now is to look more into it in order to get great results! I will keep you all in touch with this new project!

Electrophoresis Gel Results
First lane Ruler
Second lane E.coli

ASU Conference

MGE@MSA conference

Hello everyone! I hope everyone enjoyed their spring break! I know I was able to relax and clean a lot of mess I had been avoiding since the semester started! Anyway before spring break I had to chance to present my project at the MGE@MSA conference, and for being my first time attending something like this I felt pretty proud of myself! I had my poster ready and I felt ready to present it, even though I was super nervous and anxious about it. Especially when I got there and saw how great everyone’s posters were. I was impressed by the variety of research topics and the amount of students out there that have the same interests as me. It was also amazing how all of these students where from all over the world and they were excited about being here in Arizona a place that to me is no big deal.

  It was a long day but I have to admit I felt important.  I thought it was pretty neat how they provided breakfast and lunch for us and at the end of the day when we got our certificates; felt accomplished.  I know lots of my peers have been mentioning the amount of judges that passed around asking questions some had a few and others had only one but to be honest I cannot tell who my judge was! I was on the lookout for my judge but I was just not able to figure it out I know I had more than 2 people come and ask me questions some of them even gave me a few suggestions either way I was very professional and answered their questions to the best of my abilities. Like I mentioned before for being the first time I feel like I did a decent job. I enjoyed this event and I was glad to be fortunate in participating in it, and I was able to learn so many things from different people.

The speakers were all great in proving to us why going to graduate school is so important. Giving us their own life experiences as examples to how they have gained so much from this was interesting to listen to. I got to hear how graduate school opens up doors to great career opportunities that in the end result in the good life.