BME103:W930 Group2

BME 103 Fall 2012	Home People Lab Write-Up 1 Lab Write-Up 2 Lab Write-Up 3 Course Logistics For Instructors Photos Wiki Editing Help
OUR TEAM Name: Garrett Salcido Open PCR machine engineer Name: Matt Mortensen Open PCR machine engineer Name: Ramesh Tadayon Experimental protocol planner Name: Alanie Harmon Experimental protocol planner Name: Alex Torres R&D Scientist Name: Davey Rand R&D Scientist LAB 1 WRITE-UP (Please finish by 11/7/2012) Initial Machine Testing The Original Design Experimenting With the Connections When we unplugged part (part 3) from (part 6), the machine ... (did what? fill in your answer) When we unplugged the white wire that connects (part 6) to (part 2), the machine ... (did what? fill in your answer) Test Run (Write the date you first tested Open PCR and your experience(s) with the machine) Protocols Polymerase Chain Reaction How PCR Works: Step 1: Denaturation by Heat: Initially, heat separates a DNA strand into two separate strands. This is allowed because the hydrogen bonds holding the DNA together are weak and easily separable when heated. Step 2: Annealing Primer to Target Sequence: We want to target a sequence specifically and to do that you must use primers. Primers mark the ends of the target sequence. Two primers are included in the PCR one for each of the strands that were just separated during denaturation. The beginning of the DNA target sequence is also marked by the primers that bind (anneal) to the complementary sequence. Step 3: Extension: After the primers bind to the complementary DNA, the temperature is raised and the enzyme Taq DNA polymerase is used to replicate the strands. The Taq DNA polymerase, active at high temperatures, facilitates the binding and joining of the complementary nucleotides. It synthesizes an identical double-stranded DNA strand. Extension begins at the 3’ end of the primer because Taq DNA polymerase synthesizes exclusively in the 5’ to 3’ direction, so the free nucleotides are only added to the 3’ end of the primer. How to Amplify a Patient’s DNA sample 1. A PCR Machine is created to amplify a patient’s DNA sample. In order to use a PCR machine, the steps include to first gather the DNA samples, 50 μL each. 2. The frozen DNA samples are separated into tubes and once melted, 8 transfer pipettes were used to add primer. 3. Once the samples are ready, we processed the DNA in the openPCR system. Components of the PCR master mix-400μM dATP, 400μM dGTP, 400μM dCTP, 400μM dTTP and 3mM MgCl2, reaction buffer (pH 8.5), nuclease-free water Flourimeter Measurements Research and Development Specific Cancer Marker Detection - The Underlying Technology (Add a write-up of the information discussed in Week 3's class) (BONUS points: Use a program like Powerpoint, Word, Illustrator, Microsoft Paint, etc. to illustrate how primers bind to the cancer DNA template, and how Taq polymerases amplify the DNA. Screen-captures from the OpenPCR tutorial might be useful. Be sure to credit the source if you borrow images.) Results (Your group will add the results of your Fluorimeter measurements from Week 4 here)