BME100 f2013:W900 Group12 L4

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 Name: Swaroon Sridhar Name: Courtney Van Bussum Name: Christopher Lae Name: Swathi Harikumar Name: Nhi Nguyen LAB 1 WRITE-UP Initial Machine Testing The Original Design This is an image of a device called an Open PCR machine. An Open PCR Machine is a device that performs polymerase chain reactions in which DNA is modified by creating an environment that provides necessary temperature changes for this process to occur. The temperature changes consist of being heated and cooled for multiple cycles; the DNA present within the samples is separated when heated and then replicated when cooled, which in turn results in multiple copies from a single strand of DNA. The Open PCR machine can then be connected to the computer using the USB cable, which enables the user to control the number of cycles and temperatures for the experiment. With this information, the Open PCR Machine then will perform environmental changes on the DNA samples with the heating plate, heating lid, and cooling fan. Information is then processed by the circuit board, and transferred to the LED screen which displays the results. Experimenting With the Connections When we unplugged (part 3) from (part 6), the LCD display on the machine turned off and did not display anything. When we unplugged the white wire that connects (part 6) to (part 2), the machine was unable to take temperature readings. Test Run The machine was run on October 23, 2013 at 9:59AM. A major problem was encountered, in that the machine (labeled Group 12) was stuck on cycle 1 during the whole test run. (Write the date you first tested Open PCR and your experience(s) with the machine) Protocols Thermal Cycler Program Stage 1: 95°C for 3 minutes-DNA double helix unravels Stage 2: Denature; 95°C for 30 seconds-Two DNA strands separate further into two more DNA strands Anneal; 57°C for 30 seconds-Primer attaches to specific DNA sequence Extend; 72°C for 30 seconds-DNA polymerase attaches to primer and begins to add complementary nucleotides to template DNA and extend the chain Stage 3: 72°C for 3 minutes-Polymerase extends the nucleotide chain to the complementary template DNA. Target sequence begins to form. Stage 4: Final Hold; 4°C-Allows for short term storage of the reaction by stopping the thermal cycling process> DNA Sample Set-up Positive Control: Cancer DNA template Tube label: + Patient 1 ID 39526 Replicate 1 Tube Label: +1A Patient 1 ID 39526 Replicate 1 Tube Label: +1B | Patient 1 ID 39526 Replicate 1 Tube Label: +1C Negative Control: Cancer DNA template Tube label: - | Patient 2 ID 31856 Replicate 1 Tube Label: -2A | Patient 2 ID 31856 Replicate 1 Tube Label: -2B | Patient 2 ID 31856 Replicate 1 Tube Label: -2C DNA Sample Set-up Procedure Step 1 Step 2 Step 3... PCR Reaction Mix What is in the PCR reaction mix? DNA/ primer mix What is in the DNA/ primer mix? + Forward Primer + Reverse Primer Research and Development PCR - The Underlying Technology (Add a write-up, essay-style, organized into paragraphs with descriptive headers, based on the Q&A's from Section three of your worksheet) (BONUS points: Use a program like Powerpoint, Word, Illustrator, Microsoft Paint, etc. to illustrate how primers bind to the cancer DNA template, and how Taq polymerases amplify the DNA. Screen-captures from the PCR video/ tutorial might be useful. Be sure to credit the sources if you borrow images.) Polymerase Chain reaction also known as PCR is technique commonly used on laboratories in order to duplicate a certain DNA sequence and amplify the amount of DNA present. Making more DNA allows for the use of DNA in experiments, DNA profiling, research into bacterial and viral infections. The PCR reaction contiains many components which allow the complection of the reaction. Various components allow for the duplicate of DNA such as primers, templates, polymerase, magnesium, deoxyribonuleotides. Template DNA is the original DNA which is used as a template to create the complementary strands of new DNA. Due to the semi-conservative property of DNA, the new DNA strands combine to create a duplicate of the original. Primers are short pieces of DNA that are designed to match the segments of DNA to copy. They are necessary in order for DNA polymerase to attach. DNA Taq Polymerase is a naturally-occuring complex of proteins whose function is to copy a cell's DNA before it divides in two. It attaches to primers and addd nucleotides. Magnesium choloride as a calayst for the PCR reaction. Magnesium in general actas as a co-factor for DNA polymerase. The reaction cannot proceed without the presence of magnesium. Deoxyribonucleotides are the building blocks of DNA. There are four types of nucleotides designated by the different bases adenine, thymine,cytosine and guanine.