BME100 f2013:W900 Group15 L6

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BME 100 Fall 2013 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
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Name: Saiswathi Javangula
Name: Justin Dombrowski
Name: Gage Bebak
Name: Ryan Fisher
Name: abdulrahman alruwaythi
Name: student

[Instructions: add the name of your team's company and/or product here]


Computer-Aided Design


TinkerCAD is an easy tool used to create digital designs that can be 3D printed into physical objects. TinkerCAD is specifically a program that allows modeling to form or refurbish a variety of products. During the previous lab, we used TinkerCAD to improve the construction of test tubes through design techniques. During the lab, we were given PCR tubes that needed minor adjustments, which could be accurately designed through TinkerCAD. One change that our group thought to make to the PCR machine was to add more holes, or places, for the test tubes in the tray under the heating pad. By doing so, there will be more accuracy in placing the test tubes perfectly in the given compartment. Also, we made the shapes of the lids into rectangles rather than circles to properly close the lids once the process was completed. We changed the layout of the test tubes by changing the shape of the lid into rectangles, which would ultimately allow for better usage of the test tubes. This would ensure for an easier and faster method to finishing up the test tube process. By adding in rectangle lids, it will cap easier and will ensure the tips to be in place when placing them inside rather than moving around too much. Also, by squaring the lids, it would ultimately allow for more space to add labels on top of the lids. By doing so, it would be much easier to know which tube is which immediately upon looking at it.

Implications of Using TinkerCAD for Design

TinkerCAD is an excellent tool for redesigning something like the PCR machine because it becomes very easy too start with design for the PCR machine and in TinkerCAD one can easily manipulate this design by changing the size, orientation, or completely adding something. For example, one change that our group thought to make to the PCR machine was to add more holes, or places, for the test tubes in the tray under the heating pad. By doing so, there will be more accuracy in placing the test tubes perfectly in the given compartment. We could easily illustrate this change through TinkerCAD by adding in more spaces between the test tubes, along with adding more holes between the test tubes to demonstrate the change that we want implemented. Also, we added in rectangular caps rather than circular ones to allow for easier finishing of the PCR process. This could be shown through TinkerCAD by using the shapes tool to add rectangles into the area of the lids, which clearly shows what we mean to portray. Using TinkerCAD is a very easy way when trying to add more of these rows of holes into the design specs.

Feature 1: Cancer SNP-Specific Primers

Background on the cancer-associated mutation

The human body can go through a genetic mutation when cells divide for various reasons. One of the most common mutations is a single nucleotide polymorphism, or SNP for short. A polymorphism is when more than one form is expressed in a phenotype. In the case of a SNP, it means that at a certain spot of DNA from some tissue in the organism has one nucleotide while the at the same spot of DNA from the same organism but taken from a different tissue it will have a different nucleotide. The SNP rs17879961 is a SNP in the human genome located on the 22nd chromosome and actually has an impact on the human body. The gene that the rs17879961 SNP affects is the CHEK2 gene and is responsible for making the protein checkpoint kinase 2. Checkpoint kinase 2 is a protein that regulates cell division and is a tumor suppressor. If the SNP is present, the gene makes a malfunctioning protein that does not control cell division, which leads to a cancerous tumor formation.

Primer design

  • Cancer-specific Reverse Primer: GTCCTAAAAACTCTTACAC

How the primers work:

Primers are very important for the DNA amplification process. In general, the forward primer attaches to one strand of the DNA while the reverse primer attaches the complimentary strand. These primers are cancer-specific because they target the rs17879961 SNP. We can tell by looking at the last nucleotide on each primer to see if they match up(A to T, G to C). For these primers the last nucleotides do not match up because the last nucleotide on the reverse primer is the replacement nucleotide when the rs17879961 SNP is present and was designed to be that way. This causes amplification of the cancerous DNA because both strands have to be present for the DNA to copy. When non-cancerous DNA is tested with these primers, nothing happens because the reverse primer cannot attach to the non-cancerous DNA.

Feature 2: Consumables Kit

The consumables for the PCR kit which our company is creating will be tightly packed within the box that everything comes in so that it does not take up extra space and be both economically logical and will not create excessive waste. The kit will come with all necessary consumables and extras of all so that no separate purchases will be made by the consumer. It will come packaged with any liquids (including primers, etc.) on bottom so that if any leak or spill occurs there is less chance of the consumables being contaminated and as a second layer of protection, the consumables will be wrapped in plastic to reduce risk of contamination. Another goal of the way our consumables will be packaged is to have the packaging be usable to the consumer and act as a disposable tray that can hold all the pipette tips and a place in the packaging to dispose of the tips and excess liquids rather than using a trashcan or other method of disposal.

In order to address the difficulty of cleaning the slide using the micropipettor which often did not completely clean the slide, we will include small disposable napkins that can be used to easily clean the slides and simply toss the napkin into the biohazard disposal bin. The edges of the slides were extremely sharp and could be very harmful if they were to cut someone so to avoid this issue the slides in our PCR testing kit will be made out of sturdy disposable plastic. in order to avoid measuring errors the micropipettors will come with a lock which will lock the dials into place to avoid bumping them and measuring the liquids wrong and wasting the primers, etc.

Feature 3: PCR Machine Hardware

Essentially, for our system, the PCR machine will basically be included in the same way as it was in the PCR kit. However, we will be making some changes to our own PCR machine. One of the main features we want to incorporate into our PCR system is an ability to have multiple testing abilities. For example, we would want the system to analyze multiple aspects such as cancer and diabetes. To do so, we would need to add more markers. By including more markers, or primers, we could redesign the PCR machine to test multiple diseases at once. The greatest advantage to adding multiple slots in the PCR machine is that it allows patients to properly assess information all at once, making it easier to use the machine to its fullest potential.

Another function that we thought to be a major drawback for the PCR machine was the amount of time it took to perform an action. There is a relatively easy solution we would make to our own PCR machine by adding in more slots, which would mean a greater number of tubes to test a larger amount of DNA in the sample. By adding another row of holes for the test tubes or even two that could allow you to test not only 2 people at a time but 3 to 4 people. By allowing more tubes in the PCR machine, more DNA will be able to be tested, allowing more to get done in the time allocated. Therefore by adding two rows you could be essentially doubling the efficiency of the PCR device and or creating more trials to compare and thus create a larger data collection to analyze.

Feature 4: Fluorimeter Hardware

Essentially, our fluorimeter will serve the same purpose as the one used during lab. It will ultimately determine whether the sample is positive or negative for the cancer sequence. In order to fully capture the fluorescent light, a phone camera is used. The phone is placed perpendicular to the table and the camera is adjacent to the area of the drop. While in the cradle, the camera is able to face the drop and take a picture of the drop in order to determine whether the given sample is cancer positive or negative. With the entire phone layout set, a black box is then used to prevent any additional light from entering the area. When the black box is placed on top, the setting is complete to take a proper picture.

Because the biggest weakness with the fluorimeter was its inability to be reliable, the best way to fix this issue is to create a more balanced layout of the hardware. In order to do so, we changed the angle of the fluorimeter cradle so that the phone will be better able to capture the drop and not take in too much additional light. With the cradle allowing the phone to be in perfect angle adjacent to the area of the drop, the camera of the phone will be able to take a picture that will ultimately be able to distinguish whether the sample is cancer positive or negative. Because the cradle can essentially move during any given point in the experiment, the cradle will be placed in a more sturdy position to prevent any unnecessary movement. This will also help during the initial points in the lab when attempting to position the phone in a good location. Rather, by having an area that is set at a given point, time will be saved and the images will be more reliable in that most of them will be similar. Though the current layout proves to be effective, there are slight possibilities that external light may be making its way into the black box area. With the new designed version of the box, the box will be made of even thicker, black material to ensure that no light will find its way in.

Bonus Opportunity: What Bayesian Stats Imply About The BME100 Diagnostic Approach

Calculation 3 for the Bayesian statistics is P(A I B). By calculating out the probability that the patient will develop cancer given a cancer DNA sequence, we received a a vary small value that was less than 1, which ultimately states that the value is not accurate. Likewise, for calculation 4, it was the same equation; however, it asked to find the probability that the patient will develop cancer given a non-cancer DNA sequence. After our calculation, we again received a very small value that was less than one, which again proves that the value is not very accurate. Both these calculates ultimately prove that the reliability of CHEK2 PCR is not very reliable for predicting cancer. One of the biggest reasons why CHEK2 PCR was found to be unreliable could have to been due to mistakes made during lab. Some of the trials could have accidentally used more or less of the sample than required, causing for a minor error in the reading. If the CHEK2 PCR was reliable, the values for calculations 3 and 4 would have closer to 1 rather than being so small.