BME103 s2013:T900 Group5 L3: Difference between revisions
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| [[Image:BME103student.jpg|100px|thumb|Name: Cody Gates <br> Camera Operator]] | | [[Image:BME103student.jpg|100px|thumb|Name: Cody Gates <br> Camera Operator]] | ||
| [[Image:BME103student.jpg|100px|thumb|Name: Alexander Oropel<br> Research and Development Scientist]] | | [[Image:BME103student.jpg|100px|thumb|Name: Alexander Oropel<br> Research and Development Scientist]] | ||
| [[Image:BME103student.jpg|100px|thumb|Name: Matt McClintock <br> Protocol]] | | [[Image:BME103student.jpg|100px|thumb|Name: Matt McClintock <br> Data Analyzer/Protocol ]] | ||
| [[Image:BME103student.jpg|100px|thumb|Name: | | [[Image:BME103student.jpg|100px|thumb|Name: Heewon Park<br> Machine/ Device Engineering | ||
]] | |||
| [[Image:BME103student.jpg|100px|thumb|Name: Student<br>Role(s)]] | | [[Image:BME103student.jpg|100px|thumb|Name: Student<br>Role(s)]] | ||
|} | |} | ||
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| Sample Name || Ave. INTDEN* || Calculated μg/mL || Conclusion (pos/neg) | | Sample Name || Ave. INTDEN* || Calculated μg/mL || Conclusion (pos/neg) | ||
|- | |- | ||
| Positive Control || 5725984.00 || | | Positive Control || 5725984.00 || 11.66 μg/mL || N/A | ||
|- | |- | ||
| Negative Control || 2820659.67 || | | Negative Control || 2820659.67 || 4.50 μg/mL || N/A | ||
|- | |- | ||
| Tube Label: 1 Patient ID: 92336 rep 1 || 4902761.33 || | | Tube Label: 1 Patient ID: 92336 rep 1 || 4902761.33 || 9.63 μg/mL || pos | ||
|- | |- | ||
| Tube Label: 2 Patient ID: 92336 rep 2 || 4957051.00 || | | Tube Label: 2 Patient ID: 92336 rep 2 || 4957051.00 || 9.77 μg/mL || pos | ||
|- | |- | ||
| Tube Label: 3 Patient ID: 92336 rep 3 || 5446934.67 || | | Tube Label: 3 Patient ID: 92336 rep 3 || 5446934.67 || 10.93 μg/mL || pos | ||
|- | |- | ||
| Tube Label: 4 Patient ID: 44606 rep 1 || 2497338.33 || | | Tube Label: 4 Patient ID: 44606 rep 1 || 2497338.33 || 3.70 μg/mL || neg | ||
|- | |- | ||
| Tube Label: 5 Patient ID: 44606 rep 2 || 2202675.67 || | | Tube Label: 5 Patient ID: 44606 rep 2 || 2202675.67 || 2.97 μg/mL || neg | ||
|- | |- | ||
| Tube Label: 6 Patient ID: 44606 rep 3 || 1789569.00 || | | Tube Label: 6 Patient ID: 44606 rep 3 || 1789569.00 || 1.95 μg/mL || neg | ||
|} | |} | ||
<nowiki>* Ave. INTDEN = Average of ImageJ integrated density values from three Fluorimeter images</nowiki> | <nowiki>* Ave. INTDEN = Average of ImageJ integrated density values from three Fluorimeter images</nowiki> | ||
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<!--The whole team is responsible for completing this section. This is where you get to express/ argue why certain aspects fall under Must Have, Want, Must Not Have, or Should Avoid. List the two aspects that your team picked from each category in class. You are allowed to create new aspects, as long as they are relevant to your new design.--> | <!--The whole team is responsible for completing this section. This is where you get to express/ argue why certain aspects fall under Must Have, Want, Must Not Have, or Should Avoid. List the two aspects that your team picked from each category in class. You are allowed to create new aspects, as long as they are relevant to your new design.--> | ||
'''We concluded that a good system | '''We concluded that a good system "Must Have":''' | ||
* | * Results that are easy to determine. This means a clear indication of positive or negative results when compared to the controls. This is integral to the design success because the results must be easy differentiable as to not require re-testing. | ||
* | * Software that is simple to use. The software for open PCR is incredibly easy to use and a program similar would be ideal. Anything that requires computer coding or computational design is too complicated, so the software must already be made to use. A user should be able to plug in the information he or she wants and get the desired response from the software, no computing needed. | ||
'''We concluded that we would ''Want'' a good system to have:''' | '''We concluded that we would ''Want'' a good system to have:''' | ||
* | * Samples that are easily identifiable throughout the experiment. This means that there is no changing of labels or titles for the samples. This is important because it is crucial to keep the samples consistent and not mixed up. When transferring the samples there should be prepared labels to keep them straight. | ||
* | * Accessibility. This means that anyone can access the materials needed and replicate the findings. For the purpose of DNA amplification no advanced technology is required, the PCR machine is easy for anyone to access. | ||
'''We concluded that a good system ''Must Not Have'':''' | '''We concluded that a good system ''Must Not Have'':''' | ||
* | * High cost. This means that the system must not be too expensive. This is important because the system should be replicable and useful to everyone. | ||
* | * inconsistent timing. This was the most annoying problem with the original PCR design. The PCR machine kept changing times, and the group was unsure if the amplification was taking place properly. This also made it impossible to gauge the rest of the experiment timing which delayed the experiment and forced the group to reschedule further testing. | ||
'''We concluded that a good system ''Should Avoid'':''' | '''We concluded that a good system ''Should Avoid'':''' | ||
* | * High energy consumption. Possible options are a battery or solar power. Not only would this make the experiment accesible at any location, it is a more sustainable option. | ||
* | * Manual analysis of the images. This was the most tedious process of the experiment. ImageJ was useful in providing a medium for calculating light density, but a program that could do calculations on its own would be ideal. | ||
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==New System: Machine/ Device Engineering== | ==New System: Machine/ Device Engineering== | ||
Rather than consuming loads amount of energy with the PCR's technology, our new PCR will be solar battery powered. | |||
'''SYSTEM DESIGN'''<br> | '''SYSTEM DESIGN'''<br> | ||
< | http://i47.tinypic.com/65tt9e.jpg <br> | ||
http://i50.tinypic.com/35m42s8.jpg<br> | |||
< | <br>Our PCR will be powered by a removable battery that will be power by the energy provided by the sun. The solar power PCR will be not only be energy efficient, but it will also be cheaper to run, and more accessible. Using this solar power system will also give the PCR more power to run and ultimately give the new PCR more accurate and faster results. | ||
<br>'''KEY FEATURES''' | |||
'''We chose to include these new features''' | '''We chose to include these new features''' | ||
* | * Solar Battery - A removable solar battery that can be placed in the sun for hours at a time to receive energy for the PCR. This solar powered battery will create 10x more energy than a normol plug outlet can provide making the PCR, making the timing for each cycle faster and and more time consistent. The new PCR system will also have a lesser price to its users and will not consume nearly as much energy. | ||
* Plug - A area inside the PCR where the solar powered battery can be placed to turn on the PCR. This part will let the PCR be able to intake the great amount of energy that it will receive and and use the energy efficiently. | |||
'''INSTRUCTIONS''' <br> | '''INSTRUCTIONS''' <br> | ||
< | 1. Plug the solar battery into the plug<br> | ||
2. Load into PCR samples into the the PCR plate<br> | |||
3. Connect the device into a computer <br> | |||
4. Turn the PCR on<br> | |||
5. Start the computer program the PCR uses<br> | |||
<br><br> | <br><br> | ||
==New System: Protocols== | ==New System: Protocols== | ||
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<!-- If your team decided to change the PCR and/or the Fluorimeter imaging protocols, summarize the new approaches/ features here and delete the '''We chose keep the protocols the same as the original system''' section. --> | <!-- If your team decided to change the PCR and/or the Fluorimeter imaging protocols, summarize the new approaches/ features here and delete the '''We chose keep the protocols the same as the original system''' section. --> | ||
'''We chose to include these new approaches/ features''' | '''We chose to include these new approaches/ features''' | ||
* | * Plastic case - PCRs are normally found in a lab setting where safety is a big issue. Rather that having a wooden shield which is very flammable, the new shield will be be made out of plastic which will make it safer and possibly more cost efficient. | ||
* | * Solar power battery - the new battery will be more cost efficient, collect more power and will be more sustainable. | ||
* | * Plug inlet - the plug inlet will allow the solar battery plug to go into the PCR and will use the energy that is provided by the battery to be used efficiently. | ||
'''MATERIALS''' | '''MATERIALS''' | ||
Supplied by kit:<br> | |||
dNTPs<br> | |||
Reaction buffers<br> | |||
Taq DNA Polymerase<br> | |||
MgCl2 <br> | |||
<br> | |||
Supplied by Users:<br> | |||
Sample DNA<br> | |||
Forward and Reverse Primers<br> | |||
SYBR Green dye<br> | |||
Solar powered battery<br> | |||
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* '''PCR Protocol''' | * '''PCR Protocol''' | ||
Thermal Cycler Program | |||
Stage 1 | |||
95°C for 3 minutes | |||
Stage 2 | |||
35 cycles for each of the steps, each cycle will last for 30 seconds | |||
1)95°C | |||
2)57°C | |||
3)72°C | |||
Stage 3 | |||
Final Hold at 4°C | |||
* '''DNA | * '''DNA Sample Set-up Procedure''' | ||
Step 1 | |||
Insert fully charged battery in to PCR | |||
Step 2 | |||
Prepare PCR Reaction Mix and DNA sample solutions | |||
Step 3 | |||
Using a pipette, add 50μL of the DNA solutions into a labeled tube (tube should correspond with the solution) | |||
Step 4 | |||
Place tubes in the PCR | |||
Step5 | |||
Run the PCR program | |||
<br><br> | <br><br> | ||
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'''DESIGN''' | '''DESIGN''' | ||
'''Primers for PCR'''<br> | '''Primers for PCR'''<br> | ||
Amplification of cancer-associated DNA | |||
Cancer allele forward primer: 5' TATGTATGCACTGTAAGAGTT | |||
Cancer allele reverse prime: 5' CTAGGAGAGCTGGTAATTTGG | |||
A disease allele will give a PCR product because the primer associated with the process will identify the sequences that will code for cancer. From there the primer will allow for nucleotide bases to be placed in a reverse sequence from the template DNA. Essentially this will continuously amplify the cancerous DNA gene while the PCR process is in effect. | |||
<!-- If your team decided to only amplify cancer-associated DNA, list the "Cancer allele forward primer" sequence and the "Cancer allele reverse primer" sequence. Include a paragraph that explains why a disease allele will give a PCR product and the non-disease allele will not.--> | <!-- If your team decided to only amplify cancer-associated DNA, list the "Cancer allele forward primer" sequence and the "Cancer allele reverse primer" sequence. Include a paragraph that explains why a disease allele will give a PCR product and the non-disease allele will not.--> | ||
Latest revision as of 23:04, 16 April 2013
BME 103 Spring 2013 | Home People Lab Write-Up 1 Lab Write-Up 2 Lab Write-Up 3 Course Logistics For Instructors Photos Wiki Editing Help | |||||||||||||||||||||||||||||||||||||||||
OUR TEAMLAB 3 WRITE-UPOriginal System: PCR ResultsPCR Test Results
* Ave. INTDEN = Average of ImageJ integrated density values from three Fluorimeter images
Bayes Theorem is an equation in probability theory and statistics that relates inverse representations of probabilities concerning two events, or rather, it expresses a degree of change when accounting for evidence. Bayes Theorem is represented as follows: P(A|B) = P(B|A) * P(A) / P(B) Which can be read as the probability of A given B = (the probability of B given A * the probability of A) / the probability of B This information will be utilized to determine various probabilities listed below when accounting for the positive/negative values determined by the entire class as well as an outside document listing the actual yes/no cancer diagnosis
New System: Design StrategyWe concluded that a good system "Must Have":
We concluded that a good system Should Avoid:
New System: Machine/ Device EngineeringRather than consuming loads amount of energy with the PCR's technology, our new PCR will be solar battery powered. SYSTEM DESIGN http://i47.tinypic.com/65tt9e.jpg
INSTRUCTIONS
New System: ProtocolsDESIGN We chose to include these new approaches/ features
MATERIALS Supplied by kit:
Thermal Cycler Program Stage 1 95°C for 3 minutes Stage 2 35 cycles for each of the steps, each cycle will last for 30 seconds 1)95°C 2)57°C 3)72°C Stage 3 Final Hold at 4°C
Step 1 Insert fully charged battery in to PCR Step 2 Prepare PCR Reaction Mix and DNA sample solutions Step 3 Using a pipette, add 50μL of the DNA solutions into a labeled tube (tube should correspond with the solution) Step 4 Place tubes in the PCR Step5 Run the PCR program
New System: Research and DevelopmentBACKGROUND Polymerase chain reaction is the process of amplifying a strand of DNA from a DNA template strand. From here the scientist is capable of amplifying any specific gene they choose. In this research we are targeting the single nucleotide polymorphism that is rs1787996, which contains a single nucleotide variation or SNV. The CHEK2 gene is essentially a gene that is capable of coding for susceptibility to breast cancer. The relation to SNP is that it is essentially a variation of the CHEK 2 gene that is present within humans, or Homo sapiens. The cancer-related function of the gene is that it essentially changes the base Thymine to Cytosine, changing the normal allele ATT to ACT, which is the cancer related allele.
Primers for PCR Cancer allele forward primer: 5' TATGTATGCACTGTAAGAGTT Cancer allele reverse prime: 5' CTAGGAGAGCTGGTAATTTGG A disease allele will give a PCR product because the primer associated with the process will identify the sequences that will code for cancer. From there the primer will allow for nucleotide bases to be placed in a reverse sequence from the template DNA. Essentially this will continuously amplify the cancerous DNA gene while the PCR process is in effect.
New System: Software[THIS SECTION IS OPTIONAL. If your team has creative ideas for new software, and new software is a key component included in your new protocols, R&D, or machine design, you may describe it here. You will not receive bonus points, but a solid effort may raise your overall page layout points. If you decide not to propose new software, please delete this entire section, including the ==New System: Software== header.]
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