BME100 f2013:W1200 Group7 L4: Difference between revisions
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| [[Image:Carlee.jpg|100px|thumb|Name: Carlee Farhar<br>Open PCR Machine Testing]] | | [[Image:Carlee.jpg|100px|thumb|Name: Carlee Farhar<br>Open PCR Machine Testing]] | ||
| [[Image:Ambark_g7.jpg|100px|thumb|Name: Ambar Khare<br>Open PCR Machine Testing]] | | [[Image:Ambark_g7.jpg|100px|thumb|Name: Ambar Khare<br>Open PCR Machine Testing]] | ||
| [[Image: | | [[Image:batman_g7.jpg|100px|thumb|Name: Zack Silverman<br>Research and Development]] | ||
| [[Image: | | [[Image:Thalia.jpg|100px|thumb|Thalia Lebratti<br>Protocol Planning]] | ||
| [[Image:.jpg|100px|thumb| | | [[Image:MatthewCampion.jpg|100px|thumb|Name: Matthew Campion<br>Research and Development]] | ||
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=LAB 1 WRITE-UP= | =LAB 1 WRITE-UP= | ||
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'''What is a PCR?''' | '''What is a PCR?''' | ||
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A PCR machine stands for Polymerase Chain Reaction machine. When you get the PCR machine, you have to connect it to a computer via USB cable and have the OpenPCR software downloaded. This lets you in control of programming the PCR in a way you want it to work. The main function of this machine is that it uses a segment of DNA to generate billions of copies of that specific segment. DNA replication occurs naturally in the body where an enzyme known as Polymerase uses only one strand of DNA to make a complimentary strand which results in two double identical strands. A PCR however does this process repeatedly and within hours, it creates billions of copies in 35 cycles. The way it works is that the PCR goes through multiple heating cycles to produce duplicates of the segment of DNA. The very first step a PCR goes through is the process of heating up for three minutes. This is where the temperature rises up to 95 degrees C. After that, the denaturing step follows for 30 seconds which melts the DNA strand and splits the two strands apart connected by the sugar backbones. Second step is annealing for 30 seconds | A PCR machine stands for Polymerase Chain Reaction machine. When you get the PCR machine, you have to connect it to a computer via a USB cable and have the OpenPCR software downloaded. This lets you be in control of programming the PCR in a way you want it to work. The main function of this machine is that it uses a segment of DNA to generate billions of copies of that specific segment. DNA replication occurs naturally in the body where an enzyme known as Polymerase uses only one strand of DNA to make a complimentary strand which results in two double identical strands. A PCR machine however does this process repeatedly and within hours, it creates billions of copies in 35 cycles. The way it works is that the PCR machine goes through multiple heating cycles to produce duplicates of the segment of DNA. | ||
The very first step a PCR goes through is the process of heating up for three minutes. This is where the temperature rises up to 95 degrees Celsius (C) . After that, the denaturing step follows for 30 seconds which melts the DNA strand and splits the two strands apart connected by the sugar backbones. Second step is annealing for 30 seconds, this is where the temperature cools down to about 57 degrees C which then lets primers attach to the opposite ends of each strands of DNA (3' end). Because of the primers attached to the specific region of DNA, an enzyme called TAQ polymerase knows where to direct the synthesis of strands. This is known as the extending process which also lasts 30 seconds. TAQ polymerase gathers free nucleotides and binds them on each strand of DNA. The first cycle then comes to an end. The second cycle starts right away and instead of ending up with two strands, we end up with 4 strands. The next cycle we end up with 8 strands. The process keeps going on like this and we get our specific DNA segment to grow exponetially in hours. | |||
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'''PCR Design''' | '''PCR Design''' |
Latest revision as of 12:01, 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 TEAM
LAB 1 WRITE-UPInitial Machine TestingThe Original Design The very first step a PCR goes through is the process of heating up for three minutes. This is where the temperature rises up to 95 degrees Celsius (C) . After that, the denaturing step follows for 30 seconds which melts the DNA strand and splits the two strands apart connected by the sugar backbones. Second step is annealing for 30 seconds, this is where the temperature cools down to about 57 degrees C which then lets primers attach to the opposite ends of each strands of DNA (3' end). Because of the primers attached to the specific region of DNA, an enzyme called TAQ polymerase knows where to direct the synthesis of strands. This is known as the extending process which also lasts 30 seconds. TAQ polymerase gathers free nucleotides and binds them on each strand of DNA. The first cycle then comes to an end. The second cycle starts right away and instead of ending up with two strands, we end up with 4 strands. The next cycle we end up with 8 strands. The process keeps going on like this and we get our specific DNA segment to grow exponetially in hours.
Experimenting With the Connections When we unplugged the LCD display (part 3) from the circuit board (part 6), the LCD display on the machine turned off. Part 6 was the circuit board and part 3 was the LCD screen display. The circuit board is the motherboard of the PCR machine. If the wire from that is disconnected from the LCD display sceen, then the circuit board wouldn't be able to relay information to the LCD screen on what needs to be displayed. When we unplugged the white wire that connects the circuit board (part 6) to the heating plate (part 2), the temperature dropped down.
Test Run We started the test run of our PCR number 17 on Wednesday October 23rd at 12:55 p.m. After the test run, we concluded that the test run was a success and it ran exactly the way we programmed it on the software. Overall, we completed 25 of the 35 cycles which was enough to let us know if the device was working properly.
Protocols
The PCR Reaction consists of eight tubes, 50 μL, all of which contain MgCl2, dNTP, tag DNA polymerase, and the master mix. The master mix consists of 400 μM dATP, 3mM MgCl2, 400 μM dGTP, 400 μM cTTP, and 400 μM dGTP. DNA/ primer mix The DNA/primer mix consists of eight tubes, 50 μL, all of which contain a different template DNA, all of the tube contain the same reverse and forward primer.
Research and DevelopmentPCR - The Underlying Technology PCR Functions DNA Molecules
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