what is Gel Electrophoresis and how does it work?

Answer 1

An electric current moving through a saline-bathed gel separates macromolecules (DNA, proteins, etc) based on their molecular size. Smaller ones move faster than biggger ones.

Answer 2

Gel electrophoresis is a method of separating DNA, RNA, or proteins relevantly by the size of their particles. there are different ways for measuring DNA particles from RNA. If DNA measurements are involved then, a piece of fragment is cut by restriction endonucleases, which are enzymes that a DNA piece at certain sites. So you have some pieces of DNA of unknown sizes. You make a gel which serves as a molecular sponge. Then you use an electrical current to push the DNA to the other side. It makes sense that the smaller fragments would reach the other end quicker then the larger fragments. This is all analyzed by adding Ethidium Bromide which is a chemical that helps us see where these DNA particles are located within the gel, under strong UV light. This was done in the O.J. simpson case. They proved he commited the murder by putting the crime scene dna, and his dna through Gel Electrophoresis. The restriction endonuclease cut both samples at the same sites, therefore making both samples identicle.

Answer 3

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If the fragments have been run on a regular agarose gel then you could try alternative separation media. For example, there are specialized types of agarose that are much more capable of separating small fragments of double stranded DNA. Additionally, the samples could be run on a non-denaturing polyacrylamide gel where you can routinely separate fragments that differ by as little as 10 or so bp. If the samples mentioned above had been run on a lower specific concentration of agarose or polyacrylamide the next simplest thing to do is to run them on the same kind of gel but to increase the relative concentration of the separating agent. So if they did not separate on a 6% polyacrylamide gel when they were run to the bottom, you could run another sample of them on an 8% polyacrylamide gel. The higher the concentration of separating substance the greater increase in resolving power for smaller fragments.

Answer 4

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what is Gel Electrophoresis and how does it work?

Answer 5

its a separation technique, it seperates particles on basis of their size and/or charge, th promciple is simple.. u put ur sample at one end of th gel sheet & turn on an electric current particles will start moving, if th particles ur using are negativ u place them near th cathode(negative pole) so that they repel & then travel towards th anode (positive pole) & vice versa... & since they are traveling through a gel which has specific porocity (depends on th concentration of polyacrelamide for example), th distance travelled is related to th size of particle, as for charge its similar.. u can for example run a sample of diffierent length- fragments of DNA & & compare ur results with an already known control & so separate different fragments & calculate an approx. length!

i hope its useful!

Answer 6

What Is Electrophoresis

Answer 7

Gel electrophoresis is a group of techniques used by scientists to separate molecules based on physical characteristics such as size, shape, or isoelectric point. Gel electrophoresis is usually performed for analytical purposes, but may be used as a preparative technique to partially purify molecules prior to use of other methods such as mass spectrometry, PCR, cloning, DNA sequencing, or immuno-blotting for further characterization.
[edit] Separation
"Gel", refers to the matrix used to separate the molecules. In most cases the gel is a crosslinked polymer whose composition and porosity is chosen based on the weight and composition of the target of the analysis. When separating proteins or small nucleic acids (DNA, RNA, or oligonucleotides) the gel is usually made with different concentrations of acrylamide and a cross-linker, producing different sized mesh networks of polyacrylamide. When separating larger nucleic acids (greater than a few hundred bases), the preferred matrix is purified agarose (a component of agar which is a red seaweed extract). In both cases, the gel forms a solid but porous matrix that looks and feels like clear Jell-O. Acrylamide, in contrast to polyacrylamide, is a neurotoxin and needs to be handled using Good Laboratory Practices (GLP) to avoid poisoning.

"Electrophoresis", refers to the electromotive force (EMF) that is used to push or pull the molecules through the gel matrix; by placing the molecules in wells in the gel and applying an electric current, the molecules will move through the matrix at different rates, towards the anode if negatively charged or towards the cathode if positively charged (note that gel electrophoresis operates as an electrolytic cell; the anode is positive and the cathode is negative).


[edit] Visualization
After the electrophoresis runs, when the smallest molecules have almost reached the anode, the molecules in the gel can be stained to make them visible. Ethidium bromide, silver, or coomassie blue dye can be used. Other methods can also be used to visualize the separation of the mixture's components on the gel. If the analyte molecules luminesce under ultraviolet light, a photograph can be taken of the gel under ultraviolet light. If the molecules to be separated contain radioactive atoms, an autoradiogram can be recorded of the gel (as in the example shown here).

If several mixtures have initially been injected next to each other, they will run parallel in individual lanes. Depending on the number of different molecules, each lane shows separation of the components from the original mixture as one or more distinct bands, one band per component. Incomplete separation of the components can lead to overlapping bands, or to indistinguishable smears representing multiple unresolved components.

Bands in different lanes that end up at the same distance from the top contain molecules that passed through the gel with the same speed, which usually means they are approximately the same size. There are special markers available - ladders - which contain a mixture of molecules of known sizes. If such a marker was run on one lane in the gel parallel to the unknown samples, the bands observed can be compared to those of the unknown in order to determine their size. The distance a band travels is approximately inversely proportional to the logarithm of the size of the molecule.


[edit] Applications
Gel electrophoresis is used in molecular biology, genetics, microbiology and biochemistry. The results can be analyzed quantitatively by visualizing the gel with UV light and a gel imaging device. The image is recorded with a computer operated camera, and the intensity of the band or spot of interest is measured and compared against standard or markers loaded on the same gel. The measurement and analysis are mostly done with specialized software.


[edit] Nucleic acids
In the case of nucleic acids, the direction of migration, from negative to positive electrodes, is due to the natural negative charge carried on their sugar-phosphate backbone. Double-stranded DNA fragments naturally behave as long rods, so their migration through the gel is relative to their radius of gyration, or, for non-cyclic fragments, roughly size. Single-stranded DNA or RNA tend to fold up into molecules with complex shapes and migrate through the gel in a complicated manner based on their tertiary structure. Therefore, agents that disrupt the hydrogen bonds, such as sodium hydroxide or formamide, are used to renature the nucleic acids and cause them to behave as long rods again.

Gel electrophoresis of large DNA or RNA is usually done by agarose gel electrophoresis. See the "Chain termination method" page for an example of a polyacrylamide DNA sequencing gel.


[edit] Proteins
Proteins, on the other hand, can have different charges and complex shapes, therefore they may not migrate into the gel at similar rates, or at all, when placing a negative to positive EMF on the sample. Proteins therefore, are usually denatured in the presence of a detergent such as sodium dodecyl sulfate/sodium dodecyl phosphate (SDS/SDP) that coats the proteins with a negative charge. Generally, the amount of SDS bound is relative to the size of the protein (usually 1.4g SDS per gram of protein), so that the resulting denatured proteins have an overall negative charge, and all the proteins have a similar charge to mass ratio. Since denatured proteins act like they were long rods instead of having a complex tertiary shape, the rate at which the resulting SDS coated proteins migrate in the gel is relative only to its size and not its charge or shape.

Proteins are usually analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), by native gel electrophoresis, by quantitative preparative native continuous polyacrylamide gel electrophoresis (QPNC-PAGE), or by 2-D electrophoresis.


[edit] History
In a long academic book on electrophoresis edited by M Bier, [1], pages 225ff cit references from the 1800s. However, Oliver Smithies made significant contributions; Bier states "The method of Smithies ... is finding wide application because of its unique separatory power", taken in context, clearly implying Smithies' method is improvement.