Doing this online Bacterial ID Lab was very helpful, it provided a procedure of identifying bacterial samples that were given.
The Polymerase Chain Reaction Machine, better known as The PCR Machine is an expensive machine that is worth thousands of dollars. The PCR Machine provides information about DNA sequencing and is very important to this process. The work that a PCR machine conducts is altering the temperature of DNA. This type of machine takes a strand of DNA, and adds heat to the situation to in a sense "Melt" the DNA. It separates the DNA into two single strands from its double helix view. From there the machine cools the separated strands and each strand is primed. After the DNA strands have been stabilized to room temperature the primer copies a segment of DNA that is specified before hand.So overall the PCR Machine is a really important and helpful attribute to DNA sequencing, it helps in variations of defect detecting and even DNA TESTING!!
Now for The LAB!
-In this ID Lab you had to prepare DNA samples that were going to be used in the experiment. This step involved placing the bacterial colony in a microcentrifuge tube and inserting the digestive buffer. After that you have to heat the inactive digestive enzymes, which will denature and inactivate the enzymes. From there you add a counterbalance and add cell debris for removal of the sample. After these steps are completed the DNA becomes supernatant. Transfer the supernatant to the PCR tube and your prepping is done.
-The next step is to use PCR to make copies of these DNA sequences. In order for this to happen you have to add a PCR Master Mix solution for all of the tubes. To set up a positive control reaction, you must add controlled DNA to one of the tubes. However, for negative reactions you must combine de-ionized water to one of the tubes as well. After that you must insert both the tubes into the PCR Machine. The short strands that are extracted are the DNA strands that were to become of the solution from the PCR thermal cycling process.
-Continuing on from there you must purify the PCR solution. In order to do this step you have to set up the microconcentrator column and add a buffer solution to the column, then add the PCR solution to the column. The positive and negative controls will be placed in ice until they will be needed again. Then add counterbalance to the tube and put it in the centrifuge. After time has passed the DNA will have become stuck into the column and you must loosen it and put it into another tube separate from the others. Place the column into the new tube and add a buffer. From there you must add a centrifuge to be able to collect the DNA.
-The next step in this process is the PCR sequencing. First you must dilute the PCR product to increase its overall volume. Add distilled water to the new PCR Product in order to purify it. The blue and green strip tubes contain PCR reactant mixtures. Now you need to add the purified PCR solution to the mixtures. They are now ready for the PCR Amplification Process. Place the tubes into the PCR Machine. This step is now finished as well.
-Next you must load the auto sequencer and insert the tubes into it. In my first testing in this ID Lab my DNA sequence read tacggtagcgtaatgcc. Overall that completes the DNA sequencing.
