DNA Sequencing Urban Artifact's Union Terminal Beer
Posted On: 01/23/2017 - 3:40pm, Posted By: Emily Imhoff, Collections Manager - Zoology
Our Zoology curatorial team is breaking down the yeast that Urban Artifact collected from Union Terminal grounds as part of our 21+ CurioCity: Crafting Culture event happening February 9 at their location. Urban Artifact is brewing a special batch of beer using this yeast, and we want to find out what species it is! We are actually extracting the DNA from the yeast and analyzing it to determine the species. All DNA is made up of a series of nucleotides, of which there are four types: A, T, C and G. We will look at the sequence of the nucleotides of a certain region of the DNA that is known to function like a barcode that identifies different yeast species. Then we will compare the sequence of the Union Terminal yeast’s “barcode” to a database of yeast species to see if we can find a match.
First we isolate the DNA from the yeast cells. To do this, we place the yeast sample into a solution with a mild detergent and then add a special enzyme called Proteinase K. This enzyme, when heated, breaks down the cell membranes and releases the strands of DNA into the liquid.
Each tube contains a sample of yeast cells, a detergent solution, and Proteinase K enzyme. This heat block will hold the tubes at just the right temperature for the enzyme to break down the proteins in the yeast cells.
Next, we make copies of the specific fragments of the DNA that will help us identify the species of the yeast. We run each sample through a polymerase chain reaction (PCR) which involves a series of cycles of heating and cooling. First, ‘primers’ (short fragments of DNA) bind to DNA of complimentary sequence on either end of the DNA we are looking for. The reaction is then warmed up and enzymes attach to the ends of the primers and begin copying the DNA strand. Now we have 2 double stranded DNA molecules where you only had one before the reaction started. This heating/cooling is repeated over and over (up to 40 times), with the number of copies of your DNA molecule doubling every cycle 2-4-8-16-32-64…etc. After 30 cycles, you have millions of copies of your DNA fragment.
In this handy demo, each tube of red beads represents a cycle of PCR. You can see the dramatic increase as the number of DNA strands (beads) produced in each cycle doubles.
After the PCR is completed, each sample is checked to see if the reaction worked. To do this, we run them through an agarose gel and look to see if DNA fragments of the expected size are present. The gel is made of agar, a gelatin-like substance made from seaweed. If you have ever had ‘vegan gelatin,’ you may have eaten agar! Using electricity, the samples of DNA fragments are pulled through the gel. Smaller fragments move farther through the gel than larger fragments. Using UV light, we can see if we got DNA fragments of the size we expect.
Each sample that comes from the PCR machine is then loaded into a gel for “gel electrophoresis”- the process of pulling the DNA through the gel using an electrical current.
If the gel shows a successful result, we will then sequence the DNA fragments. Each sample is run through the robotic DNA sequencer machine. The machine’s computer produces a chart showing the order of the nucleobases of the DNA, which we can then compare to known sequences from different yeast species. Hopefully we will find a match, and discover what species of yeast made the brew!
The DNA sequencer reads the sequence of nucleotides in each sample, so we can compare it to known sequences of yeast species.