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Question about gel electrophoresis which is used to measure the concentration of DNA?
There is a total of 8 wells and the the two ends of the wells are not used, meaning that only 6 wells are used in this experiment. The first four wells are filled with markers while the next two wells are are filled with DNA samples. Below are the amount of markers and DNA which are filled in their respective wells (in microL):
M1 : 1
M2 : 3
M3 : 5
M4 : 10
DNA 1 : 3
DNA 2 : 5
This procedure is carried out to measure the concentration of DNA. Now, what I don't understand:
1. what is the function of the markers
2. why are four markers needed
3. how do you know how many microlitre of markers needed in each well (why 1, 3, 5 and 10)
4. why do you need two DNA samples when they're the same DNA, only different amount
5. how do you know how many microlitre of DNA to put in each well (why 3 and 5)
1 Answer
- Alone GuyLv 77 years agoFavorite Answer
You have mentioned that this procedure was performed to measure the concentration of DNA, which I am assuming is from the photoimaging of the gel.
So you have 3 marker lanes (same marker) with different amount loaded, and I do not see any point in loading 3 lanes (probably the photoimagin will involve standardizing the intensity to concenteration, which is why you have 3 lanes of, but one could also work). Another reason of 3 lanes of varying conc, can be to determine the effective conc. that would give optimal visualization (why load extra).
Since my answer to your question is photoimaging to measure concenteration, that is the only reason why you would have 2 lanes of same sample loaded, in varying amount. So now you have a graph for intensity observed for amount/concentration of DNA.
Also, as I said, you have different amount of DNA loaded for another purpose, that will be to know the optimal amount to load for your concentration so that it can be visualized (why load extra).
Source(s): I am assuming there is more explanation to the gel. - Anonymous7 years ago
I think you'll need to go back to your genetics/molecular biology instructor/lecturer/teacher/tutor with this question. I don't know enough details of the procedure you've used. I could infer that the markers are known fragments of DNA and if the unknown DNA moved through the gel by the same distance then it would be the same DNA sequence. I cannot understand why different quantities have been used.
