BME103:T130 Group 12 l2
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LAB 2 WRITE-UP
Thermal Cycler Engineering
Our re-design is based upon the Open PCR system originally designed by Josh Perfetto and Tito Jankowski.
Below is the top part of the general body of the PCR machine. It will be altered so buttons can be placed next to the LED screen.
Design changes on the top body part will include changing the layout so that it may fit input buttons that will be connected and programmed internally later. These buttons will allow an individual to set up cycling details and will remove the need of an external computer to run the device.
1. Remove the knob on the lid 2. Once the knob is removed, the bolt will be able to be removed. 3. Lower the bottom heating plate to the desired height 4. Lower the higher heating plate so the difference in space between the two plates was similar as before
For the top body part
1. Attach the part as you would normally 2. Connect the input buttons to the circuit board.
1. One pipette was used to transfer each sample into an Eppendorf tube that contained 400ml of buffer. The number of the sample used was labeled clearly and all of the sample was transferred into this Eppendorf tube.
2. Two drops of SYBR Green I was placed on the first two center drops.
3. Next, two drops of the diluted sample was added on top of the SYBR Green I solution drops.
4. The drop was then aligned with the light on the flourimeter.
5. A smartphone placed on a stand was used to take as many pictures as needed of the sample and that picture was sent to a computer for analysis.
Research and Development
Background on Disease Markers
HIV is a virus that causes A.I.D.S. (acquired immunodeficiency syndrome). HIV affects the immune system by attacking specific cells that help the immune system work. These include the T Cells that help the white blood cells fend off bacteria, disease, and viruses. A SNP related HIV is rs1024611 and it affects Chromosome 17. http://www.ncbi.nlm.nih.gov/projects/SNP/snp_ref.cgi?rs=1024611
The primer that can be used to detect HIV is ATA and its reverse TAT. A diseased allele will only give results because of the way the PCR works and how the primers will attach. If the sample has the disease then the primer will attach to the DNA and as the PCR goes through its cycles there will be a multitude of the same DNA strand with the SNP rs1024611. If the sample does not have the disease allele then there will be no interaction with the DNA and the primer, therefore there will be no change from the original and the end results.