Week 1-Day 2

Bat DNA- French and Irish bat DNA is compared in the UCD bat Lab. Changing the pH in the purification stage will result in DNA (alkaline) deposited in the aqueous phase or RNA (acidic) in the aqueous phase!

Bat DNA- French and Irish bat DNA is compared in the UCD bat Lab.
Changing the pH in the purification stage will result in DNA (alkaline) deposited in the aqueous phase or RNA (acidic) in the aqueous phase!

  • I really enjoyed today’s lab work. Clare and I again worked with Dr. Lao and followed on from the work with DNA extraction yesterday.
  • The first day ended with DNA samples extracted and purified. This involved a great deal of measuring, vortexing and centrifuging.
Very careful extraction of the alcohol, avoiding the DNA pellet.

Very careful extraction of the alcohol, avoiding the DNA pellet.

Spins at speed and separates the phases.

Spins at speed and separates the phases.

  • The lab session began with a final round of alcohol precipitation. The objective was to get a very clear, pure pellet of DNA, and I’m delighted to report that we both managed quite successfully.
  • When the DNA pellet was finally ready and nuclease free water added it is time to ascertain exactly How Much DNA is in the sample. This is done using a spectrophotometer, the Nano Drop. From the smallest quantity of solution the amount of DNA is quantified and graphed on the screen. It is tricky to place the sample into the machine and a steady hand is very useful!
Tiny Volume Needed and Steady Hand!

Tiny Volume Needed and Steady Hand!

IMG_0204 IMG_0210 Loading the Sample

  • Using the Nano drop gives you an idea if the sample contains enough DNA to continue to the next step, which is Polymerase Chain Reaction (PCR).
  • If you have a section of DNA that you have identified perhaps from one of the many data bases online, you can use PCR to produce multiple copies of a section of DNA from a single sample.
  • Although Clare and I had a very brief understanding of this process, we did spend time with Dr. Lao brushing up.  We checked out the numerous genetic data bases, how to select the primers for the experiment-NOT Following? Don’t worry we will upload a pdf and diagram version of the process.
  • Before the PCR process can begin, a buffer solution is required, this contains the primers and the enzymes-each with a unique role in the process. (pdf and diagram to follow).
Measurement is key!

Measurement is key!

IMG_0228

  • From what we learned today PCR involves heating up your DNA sample to split the double helix, cooling it down so that the primers you have added will join to the exposed strands (complementary base pairing). Then the vials are cooled and an Enzyme such as Taq Polymerase will act to Join the free nucleotides which have now using complementary base pairing attached to the exposed strands with the primers. New DNA double helix is formed.
  • The process is repeated many times-all done in the PCR machine overnight.
  • Our samples are in the PCR machine and in the morning we will run gel electrophoresis, how good are our lab skills? Tomorrow will tell! If it works you can amplify the DNA hugely.

PCR Machine

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