Lab 7B Gel Electrophoresis

Lab 7B Gel Electrophoresis Result: Screen shot of your gel 1 Analysis: (�Video) 1. What are the different kinds of molecules that can be separated using this this technique? Macromolecules like DNA, RNA and proteins. DNA fragments are separated according to their size. Proteins can be separated according to their size and their charge (different proteins have different charges) 2.What are the applications of this technique? n the separation of DNA fragments for DNA fingerprinting to investigate crime scenes. To analyze results of polymerase chain reaction Step 1. Make the Gel & Step 2: Assemble the Gel Apparatus (V lab) 3. What is buffer and what is it used for? A solution that can resist pH change upon the addition of an acidic or basic components. It is able to neutralize small amounts of added acid or base, thus maintaining the pH of the solution relatively stable. This is important for processes and/or reactions which require specific and stable pH ranges. In gel electrophoresis are used to provide ions that carry a current and to maintain the pH at a relatively constant value. 4. What is the purpose of the “comb?” Electrophoresis combs are used to create the wells in gels for electrophoresis, a technique that uses the electrical charges of molecules to separate them by their length. It is often used to analyze DNA fragments. When a gel is poured, a comb is inserted. 5. What is /are the purpose of a gel? Analysis of nucleic acids and proteins. separates molecules on the basis of their rate of movement through a gel under the influence of an electrical field. Used to determine presence and size of PCR products Step Three: Load the DNA sample into the Gel Step Three: Load the DNA sample into the Gel 5. Label the following on your bench. Your Pipette tips (F) has already been done for you. Fig 1. Gel Electrophoresis equipment A Micro pipette C DNA strand B Loading buffer D DNA size standard E Gel Bay with Buffer + Gel 6. What are the two major functions of “Loading Buffer?” 1. Make it easier to see by dying strands 2. Make them bigger 2 7. What is the function of the DNA Standard (also called the “Ladder”? Gives you a reference by which to measure the length of DNA strands. Step Four: Hook up the Electrical Current and Run the Gel 8. Since DNA has a negative charge, in which direction will it go in the gel? They move towards the positive electrode 9. If there are bubbles, what does this mean? When running agarose or polyacrylamide gels, bubbles mean the electrodes are connected, plugged in, and that current is flowing. Analysis and Conclusions: Step Five: Stain the Gel and Analyze the Results 10. What is ethidium bromide? Why can it be dangerous? It is a stain that binds to the DNA, and it can damage the DNA. 11. Read the following gel to the left. Put in the estimated base pair lengths to the left in the boxes. 12. Which fragments went the farthest through the gel, large pieces of DNA (6000 bp) or smaller pieces of DNA (1000 bp)? Smallest pieces went the farthest. 13. Gel Electrophoresis is used often in forensics. Look at the following gel to the left. From the evidence DNA, which individual matches the DNA evidence left at the crime scene? Suspect #2 matches the evidence. 3 14. When running a gel, you need to have a positive control and a negative control. What do these mean and what are they each used for? The positive control is what the DNA gravitates towards, and the negative control repels the DNA 15. Name and discuss a source of error in performing and evaluating gel electrophoresis. It is important that sources of errors in this technique be minimized in order to get accurate results. One source of error is contamination of the DNA sample. If there is foreign DNA in the sample, the gel will have more bands than would be found in a gel that contains only the purified sample 4