BME100 s2014:T Group8 L6
|BME 100 Spring 2014|| Home |
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
Wiki Editing Help
Plachecki PCR: "From Primers to Perfection"
LAB 6 WRITE-UP
TickerCAD is a computer system that can design three dimensional design images. The program is very straightforward and user friendly. Lessons teach the basics of the program and then you have the ability to create your own images. You can even import generate shapes and designs.
Our new design incorporates a computer into the PCR machine so that all data could be retrieved straight from the device wihtout having to use an external computer.
Our group noticed some aspects of Open PCR that could be improved.
What we did to change this was to attach a mini computer to the PCR. Therefore no external computer will be needed.
This computer is attached to the heater and cooler, and has a tray with dna wells that can be pulled out. The difference here is that
the wells are more in number, and are labeled as well in order to prevent mistakes.
Feature 1: Disease SNP-Specific Primers
Background on the disease-associated mutation
rs237025 is a single nucleotide polymorphism (SNP). Nucleotides are a building block for DNA, they are one of the "letters" that construct codons. Polymorphisms are a common variation in DNA sequence between individual people. The SNP rs237025 is found in homo sapiens and located on chromosome 6. It is associated with the the gene SUMO4 as well as TAB2. SUMO4 stands for small ubiquitin-like modifier 4, and is responsible for controlling target proteins' subcellular localization, stability or activity. Several diseases are linked to the SNP rs237025 such as Type 1 and 2 Diabetes, nephropathy, rheumatoid arthritis, and psoriasis vulgaris. The non-disease allele contains GTG, however a change in this allele at the "G" position is linked to the disease. The disease associated allele contains the sequence ATG.
Every PCR reaction needs two primers, one forward and one reverse. The forward primer is disease-allele-specific because it ends with the nucleotide from the disease-associated allele. The forward primer for SNP rs237025 ends with ATG, the sequence for the disease associated allele. What this means is that the forward primer will amplify DNA that contains this disease-associated SNP, and will not react to amplify DNA without the allele.
Feature 2: Consumables Kit
The consumables kit for our PCR Machine and Fluorimeter Set will include:
The light-blocking tubes for the SYBR Green address the issue of variance in the experiment due to prolonged light exposure during the experimental process. These tubes are not clear like PCR tubes and have a lockable rotational top to more easily get the substance out without over-exposing its photsensitivity.
Feature 3: Hardware - PCR Machine & Fluorimeter
The PCR machine and Fluorimeter will function the same in our new design. However, the new system will generate a more efficient process as well as make the machine easier to operate. The new design contains labeled DNA wells to avoid confusion and they are spaced slightly further apart so as to allow easier transfer using the Fluorimeter. Since the process takes a significant amount of time to complete, additional DNA wells have been added to create more efficient tests. The primary change made to redesign the PCR machine was the addition of a miniature computer on the machine itself. In the original design the PCR machine had to be plugged in to an external computer for testing, so we implemented a mini-computer to make the device more portable and reduce the chance of errors or mistakes involving an external device.
Bonus Opportunity: What Bayesian Stats Imply About The BME100 Diagnostic Approach
Calculation 3 was not very close to one. Since this is the test that estimates the probability that a patient will develop the disease when they give a positive test, this is not very reliable. Calculation 4 was a bit closer than calculation 3 to 1, but still not "very close". This calculation was for the probability that a patient will not develop the disease when they give a negative test and is slightly reliable but not the most favorable probability.