BME100 f2013:W1200 Group11 L4: Difference between revisions
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The four nucleotide bases that comprise DNA are: Adenine (A), Thymine (T), Guanine (G), and Cytosine (C. Adenine and Thymine bind to each other, | The four nucleotide bases that comprise DNA are: Adenine (A), Thymine (T), Guanine (G), and Cytosine (C. Adenine and Thymine bind to each other, | ||
and Guanine and Cytosine bind to each other. This means that a "G" nucleotide base on the template strand will bind to a "C" nucleotide base on the primer. | and Guanine and Cytosine bind to each other. This means that a "G" nucleotide base on the template strand will bind to a "C" nucleotide base on the primer. | ||
While the "A" nucleotide base on the temple strand will bind to the "T" nucleotide base on the primer. | While the "A" nucleotide base on the temple strand will bind to the "T" nucleotide base on the primer. | ||
All of these nucleotides are take by the taq polymerase in for sequencing in the DNA template. | All of these nucleotides are take by the taq polymerase in for sequencing in the DNA template. | ||
<br> | <br> | ||
''''''Diagram of PCR'''''' | ''''''Diagram of PCR'''''' |
Revision as of 15:54, 29 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 Original Design
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 the white wire that connects (part 6) to (part 2), the machine
We first tested the Open PCR machine on October 23, 2013. During this test run we had unsuccessful result, as our Open PCR machine failed to operate.
ProtocolsThermal Cycler Program 1. Denature for one cycle at 95C for three minutes (Initial hold)
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'
Thermal cycling initial starts at 95°C for three minutes during this period of time the enzymes are activated due to the optimal temperature that taq polymerase will become active. During this step the single stranded DNA template will also begin to disconnect. The next step is Denature which will be at 95°C for 30 second throughout this time period the hydrogen bonds with the complementary bases will begin to become distorted ultimately causing the DNA melting of the DNA template producing a single-stranded DNA molecule. Following denaturation, the DNA undergoes annealing, where primers form stable DNA-DNA hydrogen bonds with the template DNA. After the template DNA and the primers have bound, the DNA polymerase attaches to the primer-template hybrid. Annealing occurs at 57 degrees Celsius for 30 seconds. Then, extension occurs at 72 degrees Celsius for 30 seconds. During extension, the DNA polymerase enzyme synthesizes a new strand of DNA complementary to the template strand by adding dNTPs complementary to the bases of the template strand in the 5' to 3' direction. During the final step which occurs at 72 degrees Celsius for duration of three minutes any remaining single stranded DNA that is leftover will become fully extended. Also during the final step, the final hold occurs at four degree Celsius, which is when the entire reaction becomes stored.
'Base pairing'
Base pairing is the attachment of certain nucleotide bases to others to form a double-stranded DNA molecule.
When primers attach to the template, base pairing allows for the creation of hydrogen bonds that allow them to stick together.
The four nucleotide bases that comprise DNA are: Adenine (A), Thymine (T), Guanine (G), and Cytosine (C. Adenine and Thymine bind to each other,
and Guanine and Cytosine bind to each other. This means that a "G" nucleotide base on the template strand will bind to a "C" nucleotide base on the primer.
While the "A" nucleotide base on the temple strand will bind to the "T" nucleotide base on the primer.
All of these nucleotides are take by the taq polymerase in for sequencing in the DNA template.
'Diagram of PCR'
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