BME100 s2017:Group4 W8AM L5

BME 100 Spring 2017	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 Error creating thumbnail: Unable to save thumbnail to destination Bat-El Shabtai Error creating thumbnail: Unable to save thumbnail to destination Katie Masterson Error creating thumbnail: Unable to save thumbnail to destination Tim Styrlund Error creating thumbnail: Unable to save thumbnail to destination Mustafa Alsadiq Error creating thumbnail: Unable to save thumbnail to destination Sydney Horton Error creating thumbnail: Unable to save thumbnail to destination Name: student LAB 5 WRITE-UP PCR Reaction Report In our PCR reaction lab, our pre-lab greatly helped everyone understand each element of our task. After reviewing how and what to do, we all could easily distinguish the difference between the first and second stop on the micropipettor. For majority, our final reactions containing the DNA sample and PCR reaction solution had close amounts of the same liquid. We successfully transferred all the liquids, so none was left in the tubes. Our labeling scheme was left unchanged. Overall, our PCR reaction lab concluded to be pretty accurate and smoothly completed. Fluorimeter Procedure Imaging set-up To detect DNA we used one of group member's iPhone 7 plus. We deactivated flash to get the best to take the best picture possible. We used a box with an opening side to place the materials inside. To get a good height to take the picture we used two boxes stacked onto one another. We propped the iPhone a stand, camera facing the fluorimeter. We focused the camera and put on a 10 second timer. We shut the side of the box and waited for the image to take in darkness to get a fluorescent image. Placing Samples onto the Fluorimeter Collect all materials written in Lab A protocol. Cut empty PCR tubes in half creating two steps of four linked tubes to fit all tubes into the OpenPCR machine. Label the sides of the empty tubes with the Tune Labels using a black marker. Set tubes in rack Use the proper pipetting technique to transfer 50 µL of PCR reaction mix into this empty tube labeled as positive. Discard disposable tips into cup and refrain from re-using tips to avoid cross contamination. Use a fresh pipette to transfer the positive control DNA/ primer mix into the same tube, making the volume in the positive control PCR reaction tube 100 µL. Repeat steps 5 and 6 for the ​negative control​, ​patient 1 replicates 1, 2, and 3, and patient 2 replicates 1, 2, and 3​, all by using appropriate DNA/ primer mix for the corresponding tubes. All labeled tubes will contain 100 µL PCR reaction. Close PCR reaction tub lids very tight. Place tubes into into slots of the heating block into the PCR machine, filling all 16 slots. Start machine and wait till finished. Data Collection and Analysis Images of High, Low, and Zero Calf Thymus DNA High Calf Thymus DNA Low Calf Thymus DNA Zero Calf Thymus DNA Calibrator Mean Values Error creating thumbnail: Unable to save thumbnail to destination Calibration curves Error creating thumbnail: Unable to save thumbnail to destination Error creating thumbnail: Unable to save thumbnail to destination Images of Our PCR Negative and Positive Controls Negative control Positive Control PCR Results: PCR concentrations solved Error creating thumbnail: Unable to save thumbnail to destination PCR Results: Summary Our positive control PCR result was 28.88215284 μg/mL Our negative control PCR result was 16.06301009 μg/mL Observed results Patient 92884 : The images for patient 1 showed a lot of green under the light and were bright/white in each drop in Image J. The initial concentration resulted in 80-102 μg/mL. Patient 70752 : The images for patient 2 were glowed blue under the light and were dark/transparent when analyzed in in Image J. The initial concentration resulted in 0.6-8.9 μg/mL. Conclusions Patient 92884 : The initial concentration of DNA was very high, much higher than the positive control, concluding that patient one tested positive for the DNA. Patient 70752 : The initial concentration was very low, much lower than the negative control, concluding that patient two tested negative for the DNA. Gel Electrophoresis Error creating thumbnail: Unable to save thumbnail to destination PCR Sample Processing for Agarose Gel Electrophoresis Read table from left to right as it matches the image above respectively Lane/Well Lane 1 Lane 2 Lane 3 Lane 4 Lane 5 Lane 6 Lane 7 Lane 8 Lane 9 Sample Ladder - (Negative Control) 2-3 2-2 2-1 + (Positive Control) 1-3 1-2 1-1 Agarose gel electrophoresis is a process where charged particles pass through a gel by using an electrical current and they are separated according to their size. These particles are compared to a ladder sample that is shown on the farthest left-hand side of the image above. The results of the electrophoresis show that only the sixth lane contained a positive DNA sample while the others did not show up in the gel. They reacted just like the negative control, which was not supposed to show up in the gel. These particles move towards the cathode. This is because DNA is negatively charged so it tends to move towards the positively charged side of the gel, which is the anode.