The Original Design The Open PCR machine is a device that is used to duplicate DNA strands. The machine uses heating and cooling techniques to separate dual helix strands of DNA and replicate those strands, therefore generating new DNA. Primers, which are custom built, short pieces of DNA are placed inside the machine along with the DNA. These primers act as a landing pad for DNA polymerase, a complex and naturally-occurring protein that copies a cell's DNA then divides the double helix into two separate parts.The PCR machine completes cycles that consist of heating the existing DNA until the two strands separate, cooling the DNA so primers can attach onto the strand, and then heating up the DNA again so that DNA polymerase can copy and duplicate the strands After the PCR completes thirty to forty cycles, billions of new strands of DNA are created. .
Experimenting With the Connections
After unplugging part 3 from part 6 of the PCR machine, the LED screen shut off. Once we reconnected the wire, the light came on again.
When we unplugged the white wire that connects (part 6) to (part 2), the temperature on the screen dropped.
Test Run
The date that we first worked with Open PCR was October 22, 2013 at 9:00 am. The number of the machine our group was assigned was 17.
Protocols
Thermal Cycler Program
DNA Sample Set-up
Positive Control: Cancer DNA template Tube Lable: PC
Patient 1 ID:90616 Replicate 1 Tube Lable: 1R1
Patient 1 ID:90616 Replicate 2 Tube Lable: 1R2
Patient 1 ID:90616 Replicate 3 Tube Lable: 1R3
Negative Control: Non-Cancer DNA template Tube Lable: NC
Patient 2 ID: 91514 Replicate 1 Tube Lable: 2R1
Patient 2 ID: 91514 Replicate 2 Tube Lable: 2R2
Patient 2 ID: 91514 Replicate 3 Tube Lable: 2R3
DNA Sample Set-up Procedure
Recieve Patient ID
Clearly label each tube with the Patient and Replicate Number
Using a disposible pipette, in each tube put 50μL of the PCR Reaction Mix. Do not reuse the pipettes, get a new one for each measurement.
In each tube add 50μL of the DNA/primer mix into their respective tubes
PCR Reaction Mix
The PCR reaction mix contains taq DNA polymerase, MgCl2, and dNTP's. In each of the 8 tubes there will be 50μL of solution
DNA/ primer mix
The Dna/primer mix will contain the same forward and reverse primers in each solution, however each tube will contain a different template of DNA. Each tube will have 50μL of this 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.)
BME 100 Fall 2013
