BME103:T130 Group 5 l2: Difference between revisions
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'''PCR Protocol''' | '''PCR Protocol''' | ||
'' Create a step-by-step procedure for setting up and running PCR reactions. Your instructions should include everything from adding reagents to the tubes, to programming the PCR machine and running the reaction. | '' Kyle - Create a step-by-step procedure for setting up and running PCR reactions. Your instructions should include everything from adding reagents to the tubes, to programming the PCR machine and running the reaction. | ||
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'''DNA Measurement Protocol''' | '''DNA Measurement Protocol''' | ||
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# Collect samples generated from "PCR Protocol". | |||
# Using separate pipettes for each individual sample, transfer the 150μL into the larger test tubes containing 400 mL of the buffer solution. | |||
# Set up the fluorimeter machinery as instructed, ensuring that the system is devoid of any light, as it may prevent accurate readings. Use a "blank" sample using distilled water to ensure all machinery and processes are in order. | |||
# Using the fluorimeter equipment, add two drops of each sample onto the glass plate, followed by two drops of SYBR green. When placing the drops, one should ensure that they are initially spaced out, as they will combine when more substrate is added. | |||
#Close the system down, again preventing any light from entering the system. To record the results, a photo will be used to visually measure the presence of a positive or negative result. The fluorimeter set comes with a stand to enable a SmartPhone to be utilized. Individuals will obtain the most accurate results by setting the ISO at 800 and turning off the flash setting. | |||
# Using a different pipette for waste products, clear the sample from the glass tray, move the tray forward, and repeat with the next sample. The waste samples can be placed in a separate plastic cup, and eventually disposed of in a biohazard bin. | |||
# Repeat this process until all samples have been measured and photographed. | |||
'' | '' | ||
Revision as of 20:12, 27 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 2 WRITE-UPThermal Cycler EngineeringOur re-design is based upon the Open PCR system originally designed by Josh Perfetto and Tito Jankowski.
Key Features
Instructions
ProtocolsMaterials
PCR Protocol Kyle - Create a step-by-step procedure for setting up and running PCR reactions. Your instructions should include everything from adding reagents to the tubes, to programming the PCR machine and running the reaction.
Research and DevelopmentBayesian statistics
A=Alzheimer's B=Positive test Result [math]\displaystyle{ P(A/B)=P(B/A)P(A)/P(B) }[/math]
Background on Disease Markers Alzheimer's disease is a form of dementia that occurs with loss of brain function. It affects multiple areas of the brain associated with memory, language, personality, perception, and cognitive skills. The disease typically manifests itself through forgetfulness, but gradually progresses to inability to perform basic functions, speak, and recognize family members. Currently, there is no cure. Treatment tries to slow down the disease or at the least, manage symptoms. An SNP related to Alzheimer's disease is rs1466662 (http://www.ncbi.nlm.nih.gov/projects/SNP/snp_ref.cgi?rs=1466662). It is located on chromosome four, the intron region of NM_001142552.1 and arises from a missense mutation replacing an A with a T. It is the most significant SNP outside of the SNP linked to APOE.
The backwards primer is TAT TTT TAG AAG CGA TAA AA. The forwards primer is GCC TCT TTG CCC TCT GTT TT. An allele not containing the disease will not have the sequence that allows the primers to bind. If the primers cannot bind, then that means Taq polymerase does not know where to bind. If Taq polymerase does not bind, then the sequence does not get replicated. Therefore, there will be no PCR product. Conversely, if the disease allele is present, the primers will bind. Taq polymerase will then be able to bind to the DNA and replicate the strands, creating more double-stranded DNA yielding a PCR product.
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