BME103:T130 Group 3: Difference between revisions
Line 86: | Line 86: | ||
<font size=3>'''Fluorimeter Setup'''</font><br><br> | <font size=3>'''Fluorimeter Setup'''</font><br><br> | ||
1.) The lid was first taken off the the box and one of its sides was unbuttoned in order to create a flap.<br> | 1.) The lid was first taken off the the box and one of its sides was unbuttoned in order to create a flap.<br> | ||
2.) The box was the flipped upside down in order to create a dark environment for the camera<br> | 2.) The box was the flipped upside down in order to create a dark environment for the camera.<br> | ||
3.) A hydrophobic slide was then inserted into the flourimeter.<br> | 3.) A hydrophobic slide was then inserted into the flourimeter.<br> | ||
4.) Finally the camera phone was placed in the stand. | 4.) Finally, the camera phone was placed in the stand. | ||
<font size=3>'''Fluorimeter Measurements'''</font><br><br> | <font size=3>'''Fluorimeter Measurements'''</font><br><br> |
Revision as of 12:33, 15 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 Testing
A Polymerase Chain Reaction (PCR) Machine (shown in the above image) is used to create large quantities of specific Deoxyribose Nucleic Acid (DNA) sequences. This process consists of various heating and cooling cycles to unzip DNA strands and isolate the desired DNA strands. Experimenting With the Connections When the Liquid Crystal Display (LCD) screen is disconnected from the open PCR circuit board, the LCD screen is shut off. The circuit board provides the power and the input signals for the LCD screen. Therefore, when the two parts are not connected, the LCD screen will not function. When the 16-tube PCR block is disconnected from the PCR circuit board the block will not heat or cool. The fan and lid heater are both connected to the PCR circuit board with wires, so if this connection is disrupted, those parts will not function. Test Run The initial test was ran on October 25, 2012 on machine number 9. The DNA sample was placed into the PCR machine, closed, and activated. The machine worked well, going through all the test run cycles with relative ease. There were no problems during the test.
ProtocolsPolymerase Chain Reaction GoTaq Mix Components For 100μl reaction volume:
Patient 2 Fluorimeter Setup Fluorimeter Measurements
Image J Instructions Research and DevelopmentThe Underlying Technology Polymerase Chain Reaction, also known as PCR, is used to reproduce or amplify specific sections of DNA in large quantities. A PCR machine carries out this reaction synthetically. Components of a PCR reaction: Primer: A reagent that is artificially synthesized DNA sequence that binds specifically to the target sequence of the template DNA, in this case the sequence that is linked with cancer. If the target sequence is not present on the template DNA, then the primer will not bind and amplification will not occur. Taq Polymerase: This is an enzyme that drives DNA replication. Polymerase builds each single strand of DNA marked by a primer into a new, double-stranded DNA segment. It works by finding the ends of the primers, finding free nucleotides from an added solution, then it scans the template DNA to match the proper nucleotides and attaches these nucleotides with hydrogen bonds. The benefit of using Taq Polymerse is that it can withstand extreme temperatures and does not denature during the process of the PCR reaction. Magnesium Chloride: This compound is added to the reaction mixture and binds to the Taq Polymerase, it is used to help the reaction function smoothly and can be adjusted to control the speed of the reaction. dNTP’s: This is the mixture of free bases needed to combine to make new DNA strands that are the product of this reaction. During the thermal cycling three steps occur: This image illustrates the process described above: Specific Cancer Marker Detection The single nucleotide polymorphism (SNP), 17879961, that is being examined in this experiment is known to be linked with breast and colorectal cancer. The SNP is located on the 22nd chromosome and affects the gene checkpoint kinase 2. The sequence of this gene is: When these specific primers are placed into a sample of DNA containing the cancer marker SNP 17879961, they will bind to the complimentary base pairs in the marker. The binding of these primers can only occur if the sample DNA contains the exact marker associated with the primer, the binding will result in the amplification of the initial DNA strands. Only the DNA segments with the marker will amplify which will, after many cycles, result in having millions of copies of the segment containing the marker. Therefore, if the patient is negative for this marker, then their DNA will not be amplified and the test will be negative. The presence of amplified DNA means that the patient is positive for the cancer marker.
Equation: Conditional Probabilities ResultsPositive---------------------------------------Negative
|