BME100 f2018:Group9 T1030 L5

BME 100 Fall 2018	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: Brian Aguilar Name: Karen Ruiz Name: Krissian Hargreaves Name: Christopher Ortiz Silva Name: Tabitha Keever LAB 5 WRITE-UP PCR Reaction Report Overall, the experience was positive. We all got a better understanding of how to micropipette because of the pre-lab reading. It was specific and provided enough detail so that we could move forward. In the beginning, the first and second stop was a little confusing. It was something that we needed to get hands-on experience with sot hat we could actually feel the first and second stop. While pipetting, we were fairly confident in our abilities after multiple trials. Even so, there was a couple of tubes that ended up with liquid still in there. Because of this, the actual volumes of those samples were slightly different. Our method for labeling was constant the entire duration. Fluorimeter Procedure Imaging set-up We first got the Flourimeter box and opened the contents. We had to replace the phone stand because the original one was lower than what the experiment required. After, we got two plastic containers to prop up the device so that it was in line with the phone camera. Essentially, the phone needed to be held perpendicular to the slide and the camera needed to be parallel to the slide. Then we practiced taking a picture by putting a drop of H20 on the slide. We then closed the lid on the box and set the timer for the camera. While closing the side of the box, we covered any small cracks that light could get inside through to keep the inside dark. Placing Samples onto the Fluorimeter PLace the slides on the Flourimeter (Rough side up) Add the pipette tip and set the volume to 80 microliters Then push the plunger button to the first stop on the SYBR Green I tube Submerge the tip in the liquid and release the plunger button Transport the liquid from the tube to the slide (center lane, in between the two outer rows) by pushing down on the plunger button Then push the plunger to the first stop on the 5 micrograms/ml tube of Calf Thymus DNA Submerge the tip in the liquid and release the plunger button Transport the DNA from the tube to the same spot as the SYBR Green I Put the FLourimeter inside the box and turn on the light Repeat steps 1-9 for the 2, 1, .5, and .25 concentrations Data Collection and Analysis Images of High, Low, and Zero Calf Thymus DNA Concentration: 5 μg/mL sample Concentration: .5 μg/mL sample Concentration: 0 μg/mL sample Calibrator Mean Values Initial Concentration of 2X Calf Thymus DNA solution (micrograms/mL) Final DNA concentration in SYBR Green I solution (µg/mL) Sample Number RAWINTDEN DROP - BACKGROUND Image 1 RAWINTDEN DROP - BACKGROUND Image 2 RAWINTDEN DROP - BACKGROUND Image 3 MEAN Standard Deviation 5 2.5 C-1 6879143 6922648 6743918 6848569.67 93204.88055 2 1 C-2 4607065 4618597 4516866 4580842.67 55704.64222 1 0.5 C-3 3818519 3825758 3776186 3806821.00 26776.44859 0.5 0.25 C-4 3726002 3769837 3637980 3711273.00 67151.13322 0.25 0.125 C-5 3842724 3824868 3778035 3815209.00 33408.66491 0 0 C-6 0 0 0 0 0 Calibration curves Excel Plot 1 Excel Plot 2 Images of Our PCR Negative and Positive Controls Positive Control Negative Control PCR Results: PCR concentrations solved PCR Product TUBE LABEL MEAN (of RAWINTDEN DROP - BACKGROUND) "PCR Product Concentration (µg /mL)(Step 5 calculation) Total Dilution Initial PCR Product Concentration (µg /mL)(Step 6 calculation) G9 + 7328244.333 2.664122167 12 31.969466 G9 - 2361167.333 0.180583667 12 2.167004 G9 1-1 4327550 1.163775 12 13.9653 G9 1-2 10320315.33 4.160157667 12 49.921892 G9 1-3 7314078.667 2.657039333 12 31.884472 G9 2-1 3509394.667 0.754697333 12 9.056368 G9 2-2 2673014.333 0.336507167 12 4.038086 G9 2-3 2758167 0.3790835 12 4.549002 PCR Results: Summary Our positive control PCR result was 31.96 μg/mL Our negative control PCR result was 2.16 μg/mL Observed results Patient 17649 : High level of green flourescence Patient 28137 : There was little to no images that displayed green flourescence Conclusions Patient 17649: Positive because the concentration of the PCR DNA was high which indicates a high fluorescence observed. The similarity of the Positive control further leads us to this Conclusion Patient 28137: Slightly Positive because the concentration was barely positive but still lacked fluorescence observed.