BME100 f2013:W1200 Group3 L4: Difference between revisions
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| [[Image:Sammokdad.jpg|100px|thumb|Name: Samuel Mokdad<br>Role(s) Team Leader/ Research & Development]] | | [[Image:Sammokdad.jpg|100px|thumb|Name: Samuel Mokdad<br>Role(s) Team Leader/ Research & Development]] | ||
| [[Image: | | [[Image:Morgansucks.png|100px|thumb|Name: Morgan Jameson<br>Role(s) Initial Machine Testing]] | ||
| [[Image: | | [[Image:photo16.jpg|190px|thumb|Name: Tony Facchini<br>Role(s) Research & Development]] | ||
| [[Image: | | [[Image:IMG_0703.jpg|100px|thumb|Name: Cameron Ghods<br>Role(s) Initial Machine Testing]] | ||
| [[Image: | | [[Image:1175659_10201869462841009_654957141_n.jpg|100px|thumb|Name: Evan Epperson<br>Role(s) Protocols]] | ||
|} | |} | ||
=LAB | =LAB 4 WRITE-UP= | ||
==Initial Machine Testing== | ==Initial Machine Testing== | ||
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'''The Original Design'''<br> | '''The Original Design'''<br> | ||
[[Image: | [[Image:PCRmachine1.jpg]] | ||
The OpenPCR is a technique used to copy DNA molecules. This processs is done by dramatic tempertaure changes to isolate DNA strands from one another. To begin the machine is heated to almost boiling to denature the two strands from the original DNA. Once the two strands are seperated, the temperature decreases to allow the primers to attach and bind to complimenterary matches of DNA. This initiates the coping process and the taq polmerase then attahces (slightly higher tempertaure) to add nucleotides to the sequence to extend the strand. The process is completed with the four strands of DNA. This is the first cycle and many more occur to have billions of strands duplicated. <br> | The OpenPCR is a technique used to copy DNA molecules. This processs is done by dramatic tempertaure changes to isolate DNA strands from one another. To begin the machine is heated to almost boiling to denature the two strands from the original DNA. Once the two strands are seperated, the temperature decreases to allow the primers to attach and bind to complimenterary matches of DNA. This initiates the coping process and the taq polmerase then attahces (slightly higher tempertaure) to add nucleotides to the sequence to extend the strand. The process is completed with the four strands of DNA. This is the first cycle and many more occur to have billions of strands duplicated. | ||
The Heating plate is responsible for heating the PCR samples. | |||
The function of the LCD display is to show the current progress of the machine and ensure the users that the machine is working. | |||
The cooling fan is self explanatory, it simply cools the samples being tested. | |||
Circuit boards are the brains of the machine; they make sure everything is powered and works at the correct time. <br> | |||
'''Experimenting With the Connections'''<br> | '''Experimenting With the Connections'''<br> | ||
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| Patient 1 ID: 77842 Tube: 1A || Patient 1 ID: 77842 Tube: 1B || Patient 1 ID: 77842 Tube: 1C|| Patient 1 ID: 77842 Tube:+ Control | | Patient 1 ID: 77842 Tube: 1A || Patient 1 ID: 77842 Tube: 1B || Patient 1 ID: 77842 Tube: 1C|| Patient 1 ID: 77842 Tube:+ Control | ||
|- | |- | ||
| Patient 2 ID: 26204 Tube: 2A || Patient 2 ID: 26204 Tube: 2B || Patient 2 ID: 26204 Tube: 2C|| Patient 2 ID: 26204 Tube: - | | Patient 2 ID: 26204 Tube: 2A || Patient 2 ID: 26204 Tube: 2B || Patient 2 ID: 26204 Tube: 2C|| Patient 2 ID: 26204 Tube: - Control | ||
|} | |} | ||
'''DNA Sample Set-up Procedure''' | '''DNA Sample Set-up Procedure'''<br> | ||
Step 1: Prepare the DNA/Primer mix and the PCR Reaction mix and well as the controls (both positive and negative)<br> | |||
Step 2: Label the test tubes | |||
Step 3: Add 50μL of the DNA sample Mix to each of the correspondingly labeled reaction test tubes. *Be sure to use new pipettes each time* | |||
Step 4: Start the PCR machine and wait approximately 105 minutes while the machine goes through 35 cycles based on the set-up shown directly below.<br> | |||
HEATED LID: 100°C<br> | |||
INITIAL STEP: 95°C for 3 minutes<br> | |||
STEP: 35 cycles, denature at 95°C for 30 seconds, anneal at 57°C for 30 seconds, extend at 72°C | |||
for 30 seconds<br> | |||
FINAL STEP: 72°C for 3 minutes<br> | |||
FINAL HOLD: 4°C<br> | |||
Step 5: Analyze the results | |||
'''PCR Reaction Mix''' | '''PCR Reaction Mix''' | ||
# Taq DNA Polymerase | |||
# dNTPs | |||
# MgCl2 | |||
'''DNA/ primer mix''' | '''DNA/ primer mix''' | ||
# 3μL Forward Primer | |||
# 3μL Reverse Primer | |||
# 4μL Fluorescent Probe (specific) | |||
# 5μL Sample of each patient's DNA | |||
Latest revision as of 12:42, 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 4 WRITE-UPInitial Machine TestingThe Original Design The OpenPCR is a technique used to copy DNA molecules. This processs is done by dramatic tempertaure changes to isolate DNA strands from one another. To begin the machine is heated to almost boiling to denature the two strands from the original DNA. Once the two strands are seperated, the temperature decreases to allow the primers to attach and bind to complimenterary matches of DNA. This initiates the coping process and the taq polmerase then attahces (slightly higher tempertaure) to add nucleotides to the sequence to extend the strand. The process is completed with the four strands of DNA. This is the first cycle and many more occur to have billions of strands duplicated.
The Heating plate is responsible for heating the PCR samples.
The function of the LCD display is to show the current progress of the machine and ensure the users that the machine is working.
The cooling fan is self explanatory, it simply cools the samples being tested.
Circuit boards are the brains of the machine; they make sure everything is powered and works at the correct time. Experimenting With the Connections When we unplugged part 3 from part 6, the screen on the machine turned off.
After letting the machine run for about an hour, it was still on the first cycle. This was done on October 23,2013.
ProtocolsThermal Cycler Program
Step 3: Add 50μL of the DNA sample Mix to each of the correspondingly labeled reaction test tubes. *Be sure to use new pipettes each time* Step 4: Start the PCR machine and wait approximately 105 minutes while the machine goes through 35 cycles based on the set-up shown directly below.
Research and DevelopmentPCR - The Underlying Technology Extra Credit:
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