Transcript of DNA Electrophoresis
DNA Electrophoresis (a presentation by liz mathias) So... what the heck is DNA electrophoresis? DNA electrophoresis is a process in which strands of DNA are organized by size. (or gel electrophoresis) As you know, DNA is made up codons. Certain enzymes recognize certain codons and cut the DNA strand at that point, called a restriction site. These are called RESTRICTION ENZYMES. Everybody's DNA is unique to them. (well, except maybe twins'...) So, the exact lengths of the cut strands of DNA are also unique to each person. DNA electrophoresis is a way of organizing these cut DNA strands by length, so that DNA samples can be matched by the pattern formed by bands of same-length DNA. Why do they form bands? The stands of cut DNA are injected into an agarose gel. That's this really icky, nasty, stinky clear gel, also used in petri dishes. It's made from seaweed, which is kinda strange, but seaweed has this special unbranched polysaccharide that gels well, apparently. Longer strands of DNA experience more resistance when moving through the gel, so they don't move as far as the shorter bands in the same amount of time. Woah, woah, wait a sec. Why is the DNA moving now? Good question. Well, electrophoresis involves passing an electrical charge through the DNA in the gel. DNA is negatively charged because of phosphate groups attached to it. So when an electric charge is passed through the gel with DNA, the DNA will pass to the positively charged side. And since, like we said before, the DNA strands travel at different speeds depending on their lengths, the result is... BANDS. (seaweed) (phosphate group) (band) (codon) (At this point in the presentation, I should like to pass out the manipulatives. Feel free to manipulate at will.) WHY IS THE DNA MOVING NOW?? (here are some bands for you to ogle.) (gaze upon the codons in this strand of DNA.) (lay eyes upon these graceful strands of seaweed.) (this is why it smells weird.) (gaze gently upon these phosphate groups.) Here's the procedure for gel electrophoresis... 1) Prepare agarose gel using deionized or distilled water and agarose powder. The percent composition of agarose solution does affect the distance that the cut DNA strands will travel in the gel. Higher percentages of agarose in the solution means that more bands will be distinctly present and is therefore better for matching DNA with very small strands. Place the well comb at the 0 millimeter mark on the gel tray and pour the agarose solution so that it comes to about halfway up the teeth on the well comb. Allow the agarose to solidify for a few minutos. (well comb) (gel tray) (marvel at this wonderful image.) 2) Prepare the TBE buffer for a 1 mol/L concentration using deionized or distilled water and buffer concentrate. and away we go.... The buffer conducts the electric charge through the gel, which will be submerged during electrophoresis. 3) Place the gel tray in the electrophoresis chamber. The side with the well comb should be at the anode, or side which will be negatively charged, so that the DNA travels down the gel towards the cathode, or the side which will be positively charged. electrophoresis chamber (look at it.) 4) Pour the prepared buffer into the electrophoresis chamber so that the level of buffer is equal on both sides and the top of the gel is barely submerged. There should be enough buffer on top of the gel to cover it evenly, so that no dimples are visible around the wells, but no more than necessary. 5)Gently remove the well comb, taking care not to rip the gel. (remove.) (do not rip.) (inspect the comb-removal technique of this individual.) 6)With a micropipette, insert the DNA into the wells in the gel (just enough to fill the wells). (inserted DNA) (peer precociously at this picture.) 7)Connect the electrophoresis chamber to the machine that will generate the electric charge and allow electrocuting at a voltage of no more than 135 volts. As the DNA moves down the gel, two bands of color will form, one blue-purple and one aqua/light blue. The process is complete when the purplish band has travelled all the way down to the far end of the gel. (DNA dye beginning to separate as DNA begins to travel) (ponder these progressive images.) (purple dye) (aqua dye) (and so on and so forth.) 8)Remove the gel from the gel tray and place in a small plastic tray. Submerge the gel in final DNA dye and allow it to soak for 15- 20 minutes. Then flush the gel with deionized or distilled water or allow it to soak overnight, until the color of the gel has faded enough to allow the dyed band of DNA to be visible. (dye) (gel) (meh.) DONESIES. Just kidding. We're only done with the procedure. Oh yeah, there's more.... So why should you care about DNA electrophoresis? Well, for starters, it is kinda nifty. Other than that, there really isn't a reason. They have machines to do this stuff. But consider the following fictional scenario... MURDER. (fictional, like I said.) So let me set the scene for you... On Thursday, February 27, at 3:07 a.m. emergency dispatch received a call reporting shots fired at a warehouse in Washington, D.C. Metropolitan police officers arrived at the scene at 3:18. Upon opening the warehouse, they found 6 shipping containers, containing a total of 3000 illegal AK-47s and about 2.7 million rounds of ammunition, and the body of Petty Officer Samuel Davis, who was identified by fingerprinting. Because of the involvement of Petty Officer Davis, the Naval Criminal Investigative Service, or NCIS, was given jurisdiction for the case. The NCIS team investigating the case was headed by Special Agent Leroy Jethro Gibbs, assisted by Agents Anthony DiNozzo, Timothy McGee, and Ziva David. They processed the crime scene and took the evidence back to NCIS labs, under the supervision of forensic scientist Abby Sciuto, while the body was sent to NCIS autopsy to be examined by Medical Examiner Dr. Donald Mallard. Two separate samples of blood were taken from the crime scene, yet Ms. Sciuto was unable to use her fancy-pants machines to DNA fingerprint them because they weren't working. . Good thing Abby knows how to do DNA electrophoresis. Gibbs and the NCIS team did their investigating and discovered that Petty Officer Davis was involved in the illegal arms deal that apparently went bad. They narrowed the list of suspects down to two, neither of whom had an alibi that checked out. After some difficulty, they were taken into custody. Suspect #1 Mandy Monster- known drug dealer and money launderer. (think Natalie Portman rap) Suspect #2 Antonin Shevchenko- Ukrainian born; implicated in other illegal arms deals as well as murder, theft, and embezzlement, but never charged. Abby needed to find out if one of the blood samples matched the DNA of either of the suspects, both of whom were recently injured, perhaps at the crime scene. A positive match would place one of the suspects at the crime scene at the time of the murder. Because red blood cells do not contain DNA, Dr. Sciuto obtained a sample of DNA from the white blood cells of both blood samples, then used a PCR, or Polymerase Chain Reaction, machine to provide an adequate amount of DNA to test. (gaze upon his villainous visage.) She then prepared two gels for electrophoresis. In each, she put one sample of blood from the crime scene and DNA from both suspects and Petty Officer Davis. (At this point, I should like to place the gels upon the overhead to examine the results.) One gel revealed that one blood sample from the crime scene was from Petty Officer Davis, and the other revealed the other blood was from Antonin Shevchenko, proving that he was at the scene of the crime, despite his alibi. Full transcript
(for realsies this time) OK, so my fictional murder scenario isn't that realistic. But this basic electrophoresis is where it all began. And the fancy-pants machines still use the same basic ideas. Just... fancier. And you have to admit, it is pretty nifty. He was charged with both illegal arms trade and murder and received a life sentence in jail. Works Cited
"Agarose Gel Electrophoresis of DNA." Www.vivo.colostate.edu. Colorado State University, 15 Jan. 2000. Web. 20 May 2010. <http://www.vivo.colostate.edu>.
"DNA Electrophoresis." Wikipedia. 2010. 21 May 2010. <http://en.wikipedia.org>.
Fankhauser, David B. "Electrophoretic Separation of DNA Fragments." University of Cincinnati Clermont College, 27 Jan. 2010. Web. 20 May 2010. <http://biology.clc.uc.edu/fankhauser/labs/genetics>.
Owyoung, Palmer. "How to Separate DNA From White Blood Cells." EHow. 2010. 21 May 2010.
Solomon, Eldra P., Linda R. Berg, and Diana W. Martin. "Chapter 15: DNA Technology and
Genomes." Biology. 8th ed. USA: Thomson Brooks/ Cole, 2008. 328-31.