Week 3- DNA Extraction and SIP

During this sencond week in the lab for me I have really started to understand the whole process by which I will be using the samples I took in Astoria last week.  I have been working closely with Suzanne as she works through the stable isotope probing process on the samples she took in the Esquary in March.  I wasn't here for the earlier portions of the process but I am learning how to isolate the plasmid and check for insert and then send for sequencing at the primate center.

This is the process that I will be following to look at the characteristics of my sampled microorganisms.

• Extract DNA from sterivex tubes
• Set up SIP using cesium chloride, sulfolobus, and Ethidium Bromide and adding it to out samples.  This is the gradient.
• We then recover bands by centrifuging the tubes at 72,000 rpms for 24 hours.  
• Using SIP clean the DNA bands with EDTA and tris
• Then PCR amplification with master mix and clean PCR product.
• Then  begin cloning introducing the DNA to the vector
• Plate transform cells
• Pick a couple colonies and grow up on 2xyT
• Isolate plasmid and check for insert
• Send to the primate center for sequencing to determine what microorganism we have.

This week I have extracted the DNA, performed stable isotope probing, and did PCR amplification on the DNA on my first sample of DNA (9:40 am).  Next week my goal is to extract the DNA from all of the sterivex tubes so that I can be ready to start SIP on the other samples at the end of the week.  It has been helpful to work on different portions of the process with Suzanne on her samples because than it helps me know what I will be doing in the next few weeks on my samples I recently took.


This week on Wednesday I went to my first "Tribal Law" class because I was unable to attend the class due to my trip to Astoria on Wednesday last week.  We watched a movie showing the interviews with tribal leaders.  It was interested to learn that the tribal groups in the United States are also part of their own sovereign nations.  This means they have dual citizenship, one with their own nation and one with the United States.

On Friday, Vanessa  and Grant Law planned a fun trip to Multnomah Falls and the Bonneville Dam for the interns.  It was a pretty neat experience since I had never been to the top of Multnomah Falls.  I didn't realize how high the falls really was until I go to the top and looked down.  The Bonneville Dam was interesting.  We got to go on a tour of the generators and see how the fish ladders work to allow fish to swim through the Dam.  Our tour guide was a great resource of information.  I was a fun time but also a great time of learning about places that I had never been even having lived in Oregon my whole life.