Gel Electrophoresis
Discussion Questions
Why does DNA move through the gel matrix when electrical current is applied to the gel?
I think that they are a run because they are several errors that can be done by trying to find it and have to have extra evidence to prove the correct fact.
Why does DNA move through the gel matrix when electrical current is applied to the gel?
- It moves thru the matrix due to the fact that DNA has a negative charge and the box that it is placed in has a positive and negative charge so it attracts and makes the DNA travel down the gel.
- What factors affect the rate at which DNA fragments move through the gel?
- The size size the DNA usually affect the rate in which it travels thru the gel because if its bigger then it goes slower.
- Would you expect DNA pieces of a particular size to move faster or slower in a gel with a higher percent of agarose? Explain why.
- i think it depends on the size of the DNA because if its shorts and small then it will travel faster but its its bigger i think it will take a longer time but since agarose helps it go faster then most likely it will help it move way faster.
- Describe the steps used to analyze a gel once the electrophoresis is completed.
power is turned off the molecules stop moving and they are stuck where they stopped. The UV light exposes the groups in which the DNA and molecules are of similar size and shape. From the information they contain the scientist can access the relative size and shape of the DNA molecules they contain. - Why do you think control samples are also usually run?
I think that they are a run because they are several errors that can be done by trying to find it and have to have extra evidence to prove the correct fact.
DNA Fingerprint
1: pour restrictions enzymes into DNA
(The restrictions enzymes work like scissors)
2: Pour agarose gel into tray
(agarose acts as a molecular strainer allowing smaller pieces of DNA move thought bigger pieces)
3: pour DNA into tray
(DNA now lies in the agarose gel)
4: press power
(electrophoresis now starts DNA have a negative charge so they move foward, but smaller DNA move faster than the bigger ones and when electrophoresis ends then DNA is distributed according to their size.)
5: place nylon membrane on top gel
(the agarose gel is hard to work with then DNA is transferred to a nylon membrane )
6: Add probes to the nylon membrane in the tray
(PROBES are pieces of DNA that have been radioactively labeled. probes attached themselves to the DNA fragments on the nylon membrane, but only where their code is located )
7: place X-ray film on top of nylon membrane in tray
(the probes now are exposed on corresponding areas on the X-ray film.
8: Develop film by dragging it to the developer
The film now displays the locations on the nylon membrane where probes attached themselves to the DNA fragments (*This is the DNA fingerprint*)
9: choose the culprit
in this case Honey is the suspect of the crime and her DNA matches the ones found.
(The restrictions enzymes work like scissors)
2: Pour agarose gel into tray
(agarose acts as a molecular strainer allowing smaller pieces of DNA move thought bigger pieces)
3: pour DNA into tray
(DNA now lies in the agarose gel)
4: press power
(electrophoresis now starts DNA have a negative charge so they move foward, but smaller DNA move faster than the bigger ones and when electrophoresis ends then DNA is distributed according to their size.)
5: place nylon membrane on top gel
(the agarose gel is hard to work with then DNA is transferred to a nylon membrane )
6: Add probes to the nylon membrane in the tray
(PROBES are pieces of DNA that have been radioactively labeled. probes attached themselves to the DNA fragments on the nylon membrane, but only where their code is located )
7: place X-ray film on top of nylon membrane in tray
(the probes now are exposed on corresponding areas on the X-ray film.
8: Develop film by dragging it to the developer
The film now displays the locations on the nylon membrane where probes attached themselves to the DNA fragments (*This is the DNA fingerprint*)
9: choose the culprit
in this case Honey is the suspect of the crime and her DNA matches the ones found.
Separating Fragments of DNA by Gel Electrophoresis :
3 tubes with same DNA fragments but will be cleaved , THEY MIX DIFFERENT ENZYMES to put a different recognition system. once they have all the enzymes in the tube they will put the DNA in a gel electrophoresis which is one of the most efficient ways to separate DNA. the Negative charge of the DNA migrates down to the positive end of the electro gel the smallest fragments are the ones who travel the fastest when the DNA reaches the bottom of the gel the power then turns of. and a phosphorus light then turns on showing the results of the DA patterns. when a Researcher wants to know which DNA has the interested pattern they then copy the DNA pattern by putting it in multiple solutions and using them when needed,when the DNA is finally done and they have a pattern already figured out and set infront of them a piece of X-ray film is placed over the filter and exposes the film, revealing the locations of hybridization. From this an identical agarose gel can be created and a band interest can of be cut out of the gel.
Discussions Questions:
1. After electrophoresing DNA fragments through an agarose gel, you cut out a DNA band that is closest to the positive electrode. Does this band contain DNA fragments that are the smallest or the largest on the gel?
- They contain both but the smallest DNA fragments move faster so it contains more of those .
2. What happens to the DNA molecules when they are soaked in a basic solution?
-The DNA denatures, separating into single stranded. The basic solution is used in the blotting procedure to separate double-stranded DNA into single stranded.
3. In an experiment, you digest a linear piece of DNA with two different enzymes and then electrophorese the resulting fragments on an agarose gel. You see the following DNA bands on the gel from uncut DNA, DNA cut with restriction enzyme 1, and DNA cut with restriction enzyme 2. How many sites within the original piece of DNA does each enzyme digest the DNA?
-Enzyme 1 cuts at 1 site. Enzyme 2 cuts at 3 sites.
- They contain both but the smallest DNA fragments move faster so it contains more of those .
2. What happens to the DNA molecules when they are soaked in a basic solution?
-The DNA denatures, separating into single stranded. The basic solution is used in the blotting procedure to separate double-stranded DNA into single stranded.
3. In an experiment, you digest a linear piece of DNA with two different enzymes and then electrophorese the resulting fragments on an agarose gel. You see the following DNA bands on the gel from uncut DNA, DNA cut with restriction enzyme 1, and DNA cut with restriction enzyme 2. How many sites within the original piece of DNA does each enzyme digest the DNA?
-Enzyme 1 cuts at 1 site. Enzyme 2 cuts at 3 sites.