BME103:T130 Group 17: Difference between revisions
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4. Reaction buffers | 4. Reaction buffers | ||
'''Reagent and Volume Table''' | |||
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|'''Reagent''' || '''Volume''' | |||
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|Template DNA (20 ng) || || | |||
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|10 µM forward primer || || | |||
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|10 µM reverse primer || || | |||
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|GoTaq Master Mix || || | |||
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|dH_2O || || | |||
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|'''Total Volume'''|| || | |||
'''Description of samples:''' | '''Description of samples:''' |
Revision as of 13:05, 14 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 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
GROUP17LAB 1 WRITE-UPInitial Machine TestingThe Original Design Experimenting With the Connections When the PCB board of the LCD screen was disconnected from the PCB circuit board the display output was turned off. When the white wire connecting the 16 tube PCR block to the PCB circuit board ability to regulate the temperature of the PCR was lost.
October 25, 2012 After finishing the diagnostic analysis, the PCR was tested by setting thermal cycler program to three stages. Stage one was one cycle of 95 °C for 3 minutes, the second stage was 35 cycles, 95 °C for 30 seconds, 50 °C for 30 seconds, 72 °C for 30 seconds, stage three was one cycle of 72 °C for 3 minutes. The test run lasted for about an hour and thirty minutes and confirmed that the temperature readings on the LED of the PCR machine and the computer matched.
ProtocolsPolymerase Chain Reaction The PCR replicated the wanted DNA fragments from the patient. The PCR will heat up to 95 degree Celsius and cool down to 50°C. Then heat back up to 72°C within one cycle. Over all there will be 30 cycles. At the end of the 30 cycles we have over a billion of the wanted fragments, and 60 unwanted DNA molecule strands in the solution. Step by Step instruction to amplify the Patient's DNA Sample: 1. Need to extract the DNA from the patient. 2. Put the DNA into a special PCR tube. 3. Add primer #1 to the PCR tube with the DNA. 4. Add primer #2 to the PCR tube with DNA. 5. Add Nucleotides (the A,C,T,and G). 6. Add the DNA polymerase to the PCR tube. 7. Place the PCR tube into the thermal cycler. 8. Set the temperature of the thermal cycler to 95°C and set the machine to run 30 cycles. 9. Now the thermal cycler cools down to 50°C and primer #1 and #2 attach to the single strands of DNA. 10. Now the thermal cycler temperature changes to 72°C. This begins the DNA polymerase. This pairs the DNA with its complimentary nucleotide through to the end of the DNA strand. 11. Repeat step 8-10 29 more times. 12. During cycle #3 the wanted DNA begins to appear. 13. The wanted piece of the DNA begins to double. 14. After 30 cycles are complete over a billion wanted DNA fragments will show in the DNA solution and there will be 60 copies of unwanted DNA molecules in the solution. Components: 1. MgCl_2 2. Taq DNA polymerase that lacks 5'--> 3' 3. dNTPs 4. Reaction buffers Reagent and Volume Table
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