BME103:W930 Group9: Difference between revisions
Seth Howell (talk | contribs) |
Seth Howell (talk | contribs) |
||
Line 53: | Line 53: | ||
'''Polymerase Chain Reaction'''<br><br> | '''Polymerase Chain Reaction'''<br><br> | ||
Polymerase Chain Reaction uses thermal cycling to artificially replicate DNA in vitro. First, a piece of double stranded DNA is added to a test tube containing a solution of primers, and the enzyme DNA polymerase. This solution is heated to 95 degrees celsius in order to separate (denature) the DNA molecule. Then, the solution is cooled to 57 degrees celsius and the primers bind to the single strands of DNA (provided the base sequences match up), then the solution is heated to 72 degrees celsius and DNA polymerase adds bases to complete the new complimentary strands, doubling the amount of DNA. This is repeated several times.<br><br> | Polymerase Chain Reaction uses thermal cycling to artificially replicate DNA in vitro. First, a piece of double stranded DNA is added to a test tube containing a solution of primers, and the enzyme DNA polymerase. This solution is heated to 95 degrees celsius in order to separate (denature) the DNA molecule. Then, the solution is cooled to 57 degrees celsius and the primers bind to the single strands of DNA (provided the base sequences match up), then the solution is heated to 72 degrees celsius and DNA polymerase adds bases to complete the new complimentary strands, doubling the amount of DNA. This is repeated several times.<br><br> | ||
Procedure for amplifying a person’s DNA <br> | |||
1. add .2ng’s of a person’s dna to a tube containing the proper primers, water, and the GoTaq master mix. Repeat this step for as many replicates as you need. <br> | |||
2. create a program on the open PCR machine that will run the proper cycle for replicating DNA. <br> | |||
3. The proper program will include an initial temperature hold of 95 degrees Celsius for three minutes, followed by 35 cycles of 95 to 57 to 72 all for 30 seconds each, then a final hold for 3 minutes at 72 degrees. <br> | |||
4. Place the tubes in the open pcr machine making sure to close the lid. <br> | |||
5. run the program with the tubes inside the machine. <br> <br> | |||
'''The Master Mix'''<br><br> | '''The Master Mix'''<br><br> | ||
Line 73: | Line 81: | ||
|- | |- | ||
! Total Volume || 100.0 μL | ! Total Volume || 100.0 μL | ||
|} | |}<br><br> | ||
The eight Samples<br> | |||
The eight samples run in our pcr experiment were a positive control, a negative control, and three replicate samples from each of our patients.<br> | |||
{| border="1" style="border-collapse:collapse;" class="wikitable" | |||
|+ Patients | |||
|- | |||
! Patient ID !! Gender !! Age | |||
|- | |||
| 27150 || Female || 61 | |||
|- | |||
| 73439 || Male || 62 | |||
|}<br><br> | |||
'''Flourimeter Measurements'''<br> | '''Flourimeter Measurements'''<br> | ||
[[Image:BME103 Group9 fluorimeter.png|200px|]]<br> | [[Image:BME103 Group9 fluorimeter.png|200px|]]<br> |
Revision as of 22:46, 13 November 2012
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 TEAMLAB 1 WRITE-UPInitial Machine TestingThe Original Design The open PCR is a device that enables the splitting of DNA. It is run through many cycles of heating and cooling that enables the splitting and recombination with polomers. It connects to a computer program to run the cycles. The temperature change is controled by the heating lid and than the information is fed to the LED screen. The heating lid heats tubes that are held in the heat tube holder to the appropriate temperature and then cools them as needed in each cycle. Experimenting With the Connections When we unplugged part 3, the LCD, from part 6, the Open PCR Brains Board, the LCD on the machine stopped working and did not show anything on it. When we unplugged the white wire that connects part 6, the Open PCR Brains Board, to part 2, the heat tube, the machine no longer measures the temperature of the plate and sends it to the LCD.
First test run: 24 Oct. 2012
ProtocolsPolymerase Chain Reaction Procedure for amplifying a person’s DNA
The eight Samples
Instructions for opening images in imageJ
Research and DevelopmentSpecific Cancer Marker Detection - The Underlying Technology This report used the technique called DNA amplification. (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
|