Fire coral, symbionts, and climate change Flashcards

Question

What is symbiont shuffling?

Answer 1

Symbiont shuffling is the movement and exchange of symbiotic organisms between different host species. It allows hosts to dynamically acquire beneficial symbionts. The new symbiont strains are acquired through horizontal transfer when the coral comes into contact with free-living symbiont populations or other coral colonies hosting different strains. The symbionts are in the surrounding environment.

Answer 2

They are very stable. Colonies usually revert to their original symbionic makeup.

Answer 3

To identify symbions, and which are more prevelant in that sample. What species are residing in those fire coral colonies? Based on that, we will determine where that coral was collected - warmer shallower waters, or cooler deeper waters

Answer 4

To identify the symbiont species. They are specific to the species.

Answer 5

Getting enough copies to be visible on agarose gel AND quantifying of the DNA target

Answer 6

Denaturation, anealling, and elongation/extension

Answer 7

SYBR green. During the extension stage. More binding means more fluorescence.

Answer 8

Fluorescence increases because you have more DNA strands and more dye binding to them so it is brighter. On the graph, the lines shoots up we have enough DNA to measure fluorescence. Our line has to pass the fluorescent threshold to reveal the sample fluorescence. Any fluorescence before this is just background noise.

Answer 9

Polymerase, dNTPs, fluorescent dye, and buffer with the cofactors polymerase needs to function

Answer 10

The cycles and recording the fluorescence levels in real time. This allows you to check how much fluorescence you have after each cycle. Fluorescence plateaus at a point.

Answer 11

Figuring out the order of prevelance of the three different symbiont species in our sample to tell where in the ocean it came from - hot and high waters or cool and deep waters.

Answer 12

We could use a primer that can bind to any of the symbionts but that would not tell us how they are different; which ones are present in which fire corals and how much of it is present (quantification). That is what we want to know because that is what will tell us where it came from. And the point of that is...

Answer 13

The point at which fluorescence is above background levels. Comparatively lower ct means the sample had more DNA to start with. Which shows that that sample's DNA was higher in the coral. The lowest ct value = the highest amount of DNA = the most presence symbiot

Answer 14

When the amount of PCR doubles each cycle

Answer 15

Look for the species with the lowest ct value. This is the one with the highest abundance. Ct is the number of fluorescence required to exceed the background level - now it's valid data.

Answer 16

It is an equation that you plug your figures into x = 2 ^ (higher Ct value - lower Ct value. It gives you relative abundance as a ratio. You need at least two species present. For every one of the lower abundance symbionts, there are x of the higher abundance symbionts.

Answer 17

Different species will have DNA that denatures at different temperatures due to the difference in base sequence or length of DNA strands (shorter fragments will denature quicker). So, we raise the temperature slowly and assess how many different types of DNA there are biased on the different melting temperatures you derive. We record the temperatures at which our DNA comes apart.

Answer 18

We can see fluoresence from: - Primers binding together (primer dimers) - SYBR binding with random DNA (contaminating DNA) We need to make sure we are seeing fluorescence from the DNA we care about, not primer dimers or contaminating DNA.

Answer 19

We run a PCR test with water, not sample. This will give us a baseline level of fluorescence that comes from primer dimers and contaminating DNA (background fluorescence).

Answer 20

The peak of the graph tells you the temperature at which the sample denatures. The line that peaks at a lower temperature is our primer dimers and contaminating DNA (no template/sample peak - our background stuff). The line that peaks at a higher temperature is the sample's melting point or the temp. at which it denatures. If your graph only had one peak, there is no sample DNA in the mixture. If we are suspicious that one species DNA may not be present, we will see one peak - the two lines will denature at the same temp. This means, there is no sample in there because it matches our no template control. We are only seeing primer dimers. Remember - we want to see two different temperature peaks. One temperature peak with even two lines means there is no sample DNA. It's just our primer dimers amplifying. Use-case: Which samples contain the species DNA?

Answer 21

Irradiance is the amount of photosynthetic sunlight reaching the coral's algal symbionts. Corals adapt to different irradiance through symbiont shuffling. More depth in the ocean means less irradiance which can stress the coral. So, the coral swaps our its symbionts for a species that thrives better in deeper waters with less irradiance - symbiont shuffling.