BME100 f2013:W1200 Group11 L4: Difference between revisions
Nitish Peela (talk | contribs) |
Nitish Peela (talk | contribs) |
||
(24 intermediate revisions by 4 users not shown) | |||
Line 28: | Line 28: | ||
'''The Original Design'''<br> | '''The Original Design'''<br> | ||
A PCR machine, or Polymerase Chain Reaction machine | [[Image:1211233.png]]<br> | ||
The device displayed above is an OpenPCR Machine--a wooden box that has the ability to run Polymerase Chain Reaction and send the data to the computer for further analysis. A PCR machine, or Polymerase Chain Reaction machine is used to amplify target sequence of DNA by cycling through various extreme temperatures causing the DNA to split into two individual strand. Then small strands of nucleotides called primers inside each reaction mixture binds to the ends of each DNA molecule. Once cooling starts the polymerase attached to the ends of the primers and replicates the attached DNA. This allows the amplification of DNA. It can also be used to detect and/or diagnose certain diseases based on a person's DNA. The applicability of DNA fingerprinting includes helping forensic scientists when solving a murder (match DNA samples from suspects), and paternity testing. <br> | |||
'''Experimenting With the Connections'''<br> | '''Experimenting With the Connections'''<br> | ||
[[Image:asdlkjsda.png]]<br> | |||
Source: <nowiki>http://openwetware.org/images/7/7a/PCR_group3_.png (edited)</nowiki> | |||
When we unplugged (part 3) from (part 6), the LCD screen on the machine remained lit, but did not display any information on the screen. | When we unplugged (part 3) from (part 6), the LCD screen on the machine remained lit, but did not display any information on the screen. This is because the connection between the two was unplugged and consequently no information was sent from the Circuit board to the LCD screen. | ||
When we unplugged the white wire that connects (part 6) to (part 2), the machine was unable to take temperature readings accurately. Therefore the temperature readings were no longer displayed. | |||
'''Test Run''' | '''Test Run''' | ||
On October 23, 2013, we tested the Open PCR machine. During this test run we had unsuccessful result since our Open PCR machine failed to operate. <br> Notes: | |||
* At the start, the LED light was working and the machine seemed to be functioning | |||
* During our test, the PCR machine would not display any results | |||
* A burning smell came from the PCR machine, and was over heating | |||
* No results were displayed on the computer and the LED stopped functioning<br> | |||
==Protocols== | ==Protocols== | ||
'''Thermal Cycler Program'''<br> | '''Thermal Cycler Program'''<br> | ||
[[Image:Testrun212.png]] | [[Image:Testrun212.png]]<br> | ||
1. Denature for one cycle at 95C for three minutes (Initial hold)<br> | 1. Denature for one cycle at 95C for three minutes (Initial hold)<br> | ||
2. Run through 35 cycles of denaturing, annealing, and extending (the heating and cooling makes the DNA stronger)<br> | 2. Run through 35 cycles of denaturing, annealing, and extending (the heating and cooling makes the DNA stronger)<br> | ||
Line 84: | Line 91: | ||
'''DNA Sample Set-up Procedure''' | '''DNA Sample Set-up Procedure''' | ||
*Label each tube to reflect the controls and samples of DNA from different patients | |||
*Insert the DNA/primer mixes into the correct tubes and add PCR reaction mix to each tubes | |||
*Run OpenPCR and collect the data | |||
'''PCR Reaction Mix''' | '''PCR Reaction Mix'''<br> | ||
The pCR reaction mix is 8 tubes containing: | |||
* 50 µL of Taq DNA polymerase | |||
* 50 µL of MgCl2 | |||
* 50 µL of dNTPs | |||
'''DNA/ primer mix''' | '''DNA/ primer mix''' | ||
* | * The DNA/primer mix is 8 tubes containing 50 µL each of a different template DNA. | ||
** The primers (both forward and reverse) are the same for each sample | |||
==Research and Development== | ==Research and Development== |
Latest revision as of 11:55, 30 October 2013
BME 100 Fall 2013 | Home People 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 Photos Wiki Editing Help | |||||||||||||
OUR TEAMLAB 1 WRITE-UPInitial Machine TestingThe device displayed above is an OpenPCR Machine--a wooden box that has the ability to run Polymerase Chain Reaction and send the data to the computer for further analysis. A PCR machine, or Polymerase Chain Reaction machine is used to amplify target sequence of DNA by cycling through various extreme temperatures causing the DNA to split into two individual strand. Then small strands of nucleotides called primers inside each reaction mixture binds to the ends of each DNA molecule. Once cooling starts the polymerase attached to the ends of the primers and replicates the attached DNA. This allows the amplification of DNA. It can also be used to detect and/or diagnose certain diseases based on a person's DNA. The applicability of DNA fingerprinting includes helping forensic scientists when solving a murder (match DNA samples from suspects), and paternity testing.
When we unplugged (part 3) from (part 6), the LCD screen on the machine remained lit, but did not display any information on the screen. This is because the connection between the two was unplugged and consequently no information was sent from the Circuit board to the LCD screen. When we unplugged the white wire that connects (part 6) to (part 2), the machine was unable to take temperature readings accurately. Therefore the temperature readings were no longer displayed. Test Run On October 23, 2013, we tested the Open PCR machine. During this test run we had unsuccessful result since our Open PCR machine failed to operate.
ProtocolsThermal Cycler Program
3. Extend the DNA at 72C for 3 minutes (to stabilize the DNA) DNA Sample Set-up
DNA Sample Set-up Procedure
Research and Development'PCR - The Underlying Technology' 'Components of a PCR Reaction'
'Steps of Thermal Cycling'
'Diagram of PCR'
|