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Transcript of Gel Electrophoresis
- Proteins Gel Slab- a slab of gel which acts as a medium for the electrically induced movement of DNA between the negatively charged phosphate group of DNA and the positive electrode there can many various types of gel(such as agarose, Polyacrylamide, and Starch), but the two most common are Agarose and Polyacrylamide
The wells in the gel are made by a comb when the gel is being made. THE END Real Life Applications Video History 1930-Tiselius apparatus by Arne Tiselius
1930– sucrose gel
1940-1950- zone electrophoresis
1955– starch gels
1959 – Arylamide gels and Ornstein and Davis invented discontinuous electrophoresis
1969 – Weber and Osborn discovered denaturing agents in electrophoresis
1975 – gel electrophoresis invented by Fred Sanger
1977 – sequencing gels
late 1970s – agarose gels
1983 – pulsed field gel electrophoresis enables separation of large DNA molecules
1983 – introduction of capillary electrophoresis Agarose - the most commonly used gel created from the extract from red algae (a type of seaweed), this gel is used in horizontal electrophoresis. This is used for DNA separation to separate large molecules Used in forensics for DNA fingerprinting to find out whether a person was at the crime scene or not. Polyacrylamide- is a polymer used to create gel for electrophoresis, this is used in vertical electrophoresis. It is used for DNA and protein separation(small molecules), it is also neurotoxic. Used to identify damaged genes and genetic diseases such as sickle cell anemia. Used to find genetic similarity and kinship in organisms Power Source: used to provide an electrical current through the gel, the voltage applied depends on the size and % Agarose/polyacrylamide of the gel.
Comb- a plastic comb which is used to make the sample wells in which the DNA is placed There are also different procedures for staining in both types of gels. There are also different types of chemicals used for staining such as Ethidium Bromide and SYBR Green. Electrolyte solution: used to transfer the electrical current from the power source to the gel.
1) Put Agarose powder in flask
2) Add some liquid buffer and cover the flask loosely
3) Heat the mixture in the microwave until its one liquid mixture
4) Pour the melted mixture into the gel mold and put the plastic comb in
5) Allow the gel to solidify for a set amount of time
6) Now add some loading buffer, using the micropipette, to your DNA sample
This is a dye so the DNA will be visible later
7) Using the micropipette, draw up some of the DNA fragments which have been cleaved using a restriction endonuclease
8) Place DNA fragments in the wells
9) Connect the wires to both sides of the device. make sure the cathode is attached to the side furthest away from the wells/DNA
10)View under UV light
-the rate at which the fragments moved through the gel are inversely proportional to their size
- due to the net negative charge on the phosphate groups of the DNA backbone, the fragments move towards the positive electrode(cathode) Sources Raven Johnson Losos Singer Biology textbook 7th edition
https://sites.google.com/a/luther.edu/genetics/students/claire-addis/gel-electrophoresis Reading Results
-the further the fragments are from their starting well, the shorter they are
-the larger the fragment the slower the rate
-the fragments measured by Bp(base pairs) which can also have prefixes such as Kilo and Mega