BME103:T130 Group 1 l2: Difference between revisions
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'''PCR Protocol''' | '''PCR Protocol''' | ||
Instructions: | |||
1.) Place the PCR machine in a leveled surface and plug into an outlet. | |||
2.) Collect a sample of DNA, such as blood sample. | |||
3.) Add primers to the DNA sample along with the magnesium chloride buffer and the base. | |||
4.) Place the mixed solution into the machine. | |||
5.) turn the machine up to 95 degree Celsius to separate the double stranded DNA for 3 minutes. | |||
6.) Then decrease the temperature to 57 degree Celsius for 30 seconds for the primers to bind to the separated DNA strand, forming the | |||
new double stranded DNA. | |||
7.) Then turn the temperature up to 72 degree Celsius for polymerization. | |||
8.) Repeat the temperature adjustment cycle for 34-35 cycles. | |||
'''DNA Measurement Protocol''' | |||
1.) After you add SYBR Green to the duplicated DNA sample, place the resulting DNA strand into a fluorimeter. | |||
2.) Shine light onto the sample and use a photo-taking device to record the picture of the glowing sample. | |||
3.) Use imagej to calculate the INTDEN. | |||
4.) Redo the process for a control group sample, in which the sample does not contain the DNA strand or segment that causes a certain | |||
disease. | |||
==Research and Development== | ==Research and Development== |
Revision as of 12:54, 28 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.
ProtocolsMaterials
Instructions: 1.) Place the PCR machine in a leveled surface and plug into an outlet. 2.) Collect a sample of DNA, such as blood sample. 3.) Add primers to the DNA sample along with the magnesium chloride buffer and the base. 4.) Place the mixed solution into the machine. 5.) turn the machine up to 95 degree Celsius to separate the double stranded DNA for 3 minutes. 6.) Then decrease the temperature to 57 degree Celsius for 30 seconds for the primers to bind to the separated DNA strand, forming the new double stranded DNA. 7.) Then turn the temperature up to 72 degree Celsius for polymerization. 8.) Repeat the temperature adjustment cycle for 34-35 cycles.
1.) After you add SYBR Green to the duplicated DNA sample, place the resulting DNA strand into a fluorimeter. 2.) Shine light onto the sample and use a photo-taking device to record the picture of the glowing sample. 3.) Use imagej to calculate the INTDEN. 4.) Redo the process for a control group sample, in which the sample does not contain the DNA strand or segment that causes a certain disease. Research and DevelopmentBackground on Disease Markers The disease marker being used is SNP (single nucleotide polymorphism) rs35685286. This is the marker for sickle-cell disease found on chromosome 11. Patients with sickle-cell disease have red blood cells that are mishapen and are a "sickled" or crescent shape. This results in less oxygen being carried to the patient's body tissues. Therefore, patients experience crisis, where they have severe pain in their bones in their backs or chest. These symptoms can last for hours or even days. More information about this particular SNP can be found at: http://www.ncbi.nlm.nih.gov/projects/SNP/snp_ref.cgi?rs=35685286
Step A shows the PCR heating up to 95 degrees Celcius so the hydrogen bonds between the base pairs of the double stranded DNA can be melted and separated. Next in step B, the machine is cooled to 50 degrees Celcius where the reverse primers come in and attach to the single DNA strands to create two new double stranded DNA. Finally in step C, the PCR machine is heated up to 72 degrees Celcius and the strands are fully replicated and the process repeats for approximately 30 cycles. |