BME100 s2014:W Group13 L4

BME 100 Spring 2014	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 Name: Nicole Phillips Role: Leader, Experimental Protocol Professional Name: Francis Taguinod Role:Research and Development Adept Name: Christian "Chin-Chin" Kaiser Role: PCR Machine Specialist Name: Augusta Dumanski Role: Team Mascot, Experimental Technician Name: Christiana Montoi-Reeves Role: Research and Development Guru LAB 1 WRITE-UP Initial Machine Testing The Original Design The OpenPCR machine is an affordable device which uses the process of PCR (Polymerase Chain Reaction) to read the sequences of DNA. This is done in three steps: 1. The DNA is seperated into two strands from its normal double helix shape in order to make it readable by heating the DNA up to 95 degrees celcius in the PCR heat block. 2. Primers (strands of DNA which contain the complementary sequence to the start and end points of the strand we want) bind to the strands. This is facilitated by the heat block dropping to a temperature of 57 degrees celcius. 3. The DNA polymerase attaches to the strand at the starting point primer and creates a complementary strand until it falls off of the end of the DNA strand. The PCR machine facilitates this by heating the heat block back up to 72 degrees celcius. The machine does thirty five rounds of this, which creates over a billion fragments of the target sequence. Experimenting With the Connections When we unplugged (part 3) from (part 6), the LED display on the machine turned off. When we unplugged the white wire that connects (part 6) to (part 2), the machine's temperature dropped significantly. Test Run The date we tested the machine on was March 19, 2014. We accidentally unplugged the machine and it messed it up. We ran it again and it worked fine. Protocols Thermal Cycler Program DNA Sample Set-up TAGCON+ TAG11 TAG12 TAG13 TAGCON- TAG21 TAG22 TAG23 DNA Sample Set-up Procedure Obtain necessary materials (DNA mix, PCR mix, tubes, pipette, gloves) Label tubes. Use the pipette to transfer the PCR mix and DNA mix into the tubes. Place tubes inside of PCR machine. Start machine. When machine finishes cycles, record results. PCR Reaction Mix The PCR reaction mix contains taq DNA polymerase, MgCl2, and dNTP's. DNA/ primer mix The DNA/ primer mix contains a DNA sample, a primer and a reverse primer. Research and Development PCR - The Underlying Technology Question 1) The function of each PCR reactions are as follows. The template DNA is the base of the replication of the DNA, without it the DNA cannot be amplified by itself so it is an essential part of the PCR reaction. The primers are used to bind the DNA at the origin to start the replications. The Taq Polymerase is used to create a complementary strand of the DNA strands. The dNTP's (Deoxyribonucleotides) are the building blocks of the ordinary strands. Question 2) Thermal Cycling The Denature step involves seperating the two DNA strands from each other at the temperature of 95 degrees celsius The Anneal step involves the primers being attached to the DNA strands The extend step involves the polymerase attaching to the strand of the end of the primer The final step involves the polymerase creating a complementary strand of the DNA Question 3) Each of the DNA nucleotides are paired with only one type, the Adenine pairs with the Thymine and vice versa while the Guanine pairs with the Cytosine and vice versa.