BME100 f2013:W900 Group14 L4

BME 100 Fall 2013	Home People Lab Write-Up 1 | Lab Write-Up 2 | Lab Write-Up 3 Lab Write-Up 4 | Lab Write-Up 5 | Lab Write-Up 6 Course Logistics For Instructors Photos Wiki Editing Help
OUR TEAM Name: Rachael Brard Role(s) Name: Andrew Cable Role(s) Name: Austin Davis Role(s) Name: Omar Eltohamy Role(s) LAB 1 WRITE-UP Initial Machine Testing The Original Design http://openwetware.org/images/thumb/e/ef/HALEY.png/562px-HALEY.png http://www.wired.com/reviews/wp-content/uploads/2011/12/reviews_pcr_f.jpg PCR stands for polymerase chain reaction. The PCR machine depicted here is a biochemical technology that when used properly can amplify a selected piece of DNA to make millions of copies. This machine heats DNA to break it apart, then cools it so that the polymerase will attach to the DNA strands and make copies. For the heating process, the Open PCR machine is built with highly thermally conductive alloy of aluminum and insulation, allowing heat to be kept in the machine as well as spread the temperature across the block and be able to affect the test tubes. With a adjustable lid on top of the machine, condensation of water in the test tubes is essentially negated, allowing correct and reproducible results to appear. Last but not least, the Open PCR machine has the ability to connect to essentially any computer, compatible with a Windows or Mac though USB. Also the GUI program is only necessary for the initial and final steps, allowing a disconnection from the PC. The LED machine indicates the status of reaction, such as temperature, cycles, as well as remaining time until the reaction is complete. Finally, due to a file system interface via USB, the Open PCR machine can integrate into lab or other external control systems. Experimenting With the Connections When we unplugged (part 3) from (part 6), the machine's screen went black. The LCD screen did not receive power while the rest of the machine still did. When we unplugged the white wire that connects (part 6) to (part 2), the machine did not read temperature properly. It briefly fluctuating upon detaching the wire, then read -40 degrees C. Test Run Wednesday, October 23, 2013 During our test run, we had some difficulty starting up the machine, as it had troubles starting up the process, and was constantly changing the time needed to create the first batch. Protocols Thermal Cycler Program 1. One cycle: 95 degrees Celsius for 3 minutes. 2. 35 cycles: Denature at 95 degrees Celsius for 30s, anneal at 57 degrees Celsius for 30s, extend at 72 degrees Celsius for 30s. 3. One cycle: 72 degrees Celsius for 3 minutes. 4. Hold: 4 degrees Celsius. DNA Sample Set-up row 1 cell 1 row 1 cell 2 row 1 cell 3 row 1 cell 4 row 2 cell 1 row 2 cell 2 row 2 cell 3 row 2 cell 4 DNA Sample Set-up Procedure First, the sample of DNA to be amplified is added. Both DNA primers are then added in succession. DNA Taq Polymerase is then added to the vials. Finally, loose nucleic acids are added so as to form the DNA copies. Place the tubes with the contents inside the Open PCR machine. Ensure that the lid is closed and everything is properly connected. Run Open PCR Gather tubes and use store them for use in the future. Clean up when done, ensuring all materials are accounted for. PCR Reaction Mix The PCR reaction mix is a combination of DNA to be amplified, two DNA primers, which allow for DNA Taq Polymerase to read and copy the desired sequence, DNA Taq Polymerase, for copying and reading the desired DNA, and nucleic acids, which are used to form the new copies of DNA. DNA/ primer mix What is in the DNA/ primer mix? Research and Development PCR - The Underlying Technology (Add a write-up, essay-style, organized into paragrpahs with descriptive headers, based on the Q&A's from Section three of your worksheet) (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 PCR video/ tutorial might be useful. Be sure to credit the sources if you borrow images.)