Week 3 - First Things FIrst

Here is some preliminary data that I obtained this past week. It quantifies the concentration and purity of the DNA material that I extracted from the S. aureus bacteria using two different protocols out of the six that I'll be testing. One method was the Boiling Protocol (BP) while the other one was Lysis Protocol (LP). Note the LP data unlike the BP data  has only two rows of data instead of three, and I'll tell you why in just minute.

Sample ID	Date and Time	Nucleic Acid Concentration	Unit	260/280	Sample Type	Factor
BP1	3/2/2017 4:03:11 PM	86.8	ng/µl	1.42	DNA	50
BP2	3/2/2017 4:06:24 PM	83	ng/µl	1.44	DNA	50
BP3	3/3/2017 9:18:44 AM	75.5	ng/µl	1.36	DNA	50
LP1	3/2/2017 4:52:43 PM	82.9	ng/µl	1.58	DNA	50
LP2	3/2/2017 4:54:43 PM	93.7	ng/µl	1.6	DNA	50
LP3	not analyzed as of 3/3/2017	n/a	n/a	n/a	DNA	n/a

But first let me explain why this is my starting point for my series of experiments this semester.

Out of the six protocols that are going to be tested to determine which one works best to extract more and better quality DNA material from the S. aureus bacteria there are three protocols that require an "air-dry" step at some point in their list of processes. Those protocols that require that step are: the Boiling Protocol, the Lysis Protocol, and the Meat Tenderizer Protocol; the rest of the protocols do not require that step.

In order to have a "fair competition" among all six protocols, I figured that it would be important to determine the best way to go about this air-dry" step and so I developed the following question:

Question: What is the length of time that a sample should air-dry for in order to obtain the highest and most pure DNA material yields?
Hypothesis: Allowing a sample a period of 24 hours should be enough time for a sample to completely air-dry and render maximum quality yields.

Now before I contrast the data above I should mention the following:
-The Meat Tenderizer Protocol (MTP) is a continuation of the Lysis Protocol, therefore, it's results will be those of the LP run.
-As I was swirling the sample for the LP2 run , I spilled some of the sample on a prior to adding the buffer and I attribute the gap difference in the nucleic acid concentration between LP1 and LP2 to that mishap.
-I ran out lab time and I had to modify my experiment as detailed below.
Initially:
     I was going to allow 30 minutes, 2 hours, and 24 hours of air-dry time then take the next step and compare the nuclei acid concentrations and 260/280 purity value in the end.
What actually happened:
     Because I  allowed 30 minutes of air-dry time, which was insufficient, and because the laboratory hours where coming to a close I decided to expose the liquid samples to a hot plate to speed up the vaporization of the liquid mixture on 2 out of the 3 samples per protocol to move on to the next step. I figured I would compare the values that I was testing of the samples that where exposed to heat vs the sample that was given 24 hours (plenty of time, so I thought) to air-dry and determine if the exposure to the heat affected the yields or quality.
     Once dry, I added the respective buffers and measured the values of interest using a spectrophotometer apparatus (nano-drop machine) and the results are on the above table over the first two rows for each different method (BP1, BP2, and LP1 and LP2). These where expected to be similar because, minus to accidental spill, they were put through the same processes under the same conditions; and so they are relatively close values.
     The morning after, I came back to the lab to check on the samples labeled #3 of both protocols, and I was surprised to see that one of them still hadn't evaporated completely and that was LP3. BP3 had completely evaporated and was air-dry which is what the protocol asks for. What I did next was suspend it in its buffer and analyze the content using the spectrophotometer. The results are those on the third row on the first set of results. The LP3 sample was still liquid so I chose to let it air dry over the weekend and measure the values of interest next week.
Results:
     As far as the Boiling Protocol, upon comparing the concentration and quality of the DNA from the sample that was allowed to completely air dry for 24 hours on its own versus the two samples that where heated dry in less than 30 minutes the data shows that there is more quantity and quality if this step is accelerated by heat drying the liquid mixture. So, by heating you gain time and you end up having more and better quality product.
     In regards to the Lysis Protocol, I have to wait until I get back into the lab to finish the protocol for the third sample and then compare the data to determine what the best method will be to yield higher and better nucleic acid.