BME100 f2015:Group10 1030amL5

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OUR TEAM

JOEL HARNISCH
GERRIT ORTHLIEB
BRANDON BURGERS
SAI ADONI
SERGIO MEJIA


LAB 5 WRITE-UP

PCR Reaction Report

Our team conducted the procedure of pipetting the way it was described in the pre-lab readings and watching video modules helped with getting familiar on how to use a pipette. Each team member understood the difference between the first and second stop on the pipettor. The first stop on the pipettor is used to collect a sample once finger has been removed from stop and sample has been collected. The second stop is used to release all of the sample from the pipettor into the desired location. The final reactions had the same amount of liquids for all tubes because the pipettor was calibrated to intake a volume of 120 micro-liters. Sample of DNA and PCR reaction mix were left over because the procedure was focused on taking a specific amount of sample for PCR reaction. Prior labeling of tubes from previous investigation worked efficiently for labeling tubes and avoiding cross contamination for this investigation. Pipetting was done successfully in the group by following the pre-lab reading steps and instructions on day of the lab.

Fluorimeter Procedure

Brandon: Smart Phone Camera Settings

  • Type of Smartphone: iPhone 6
    • Flash: No flash
    • ISO setting: 800
    • White Balance: Auto
    • Exposure: 100%
    • Saturation: 100%
    • Contrast: 0%


Camera set-up
The smartphone must be set camera and placed in a cradle at distance such that the image will not be blurred and the camera can focus. The camera must be perpendicular to the drop on the slide. The distance between the cradle and fluorimeter was about 7 centimeters. At this distance, we could capture a clear, accurate image of the samples in the fluorimeter.

  • Distance between the smart phone cradle and drop =

7

IF THIS FOLLOWING PORTION IS BLANK PLEASE GIVE BRANDON A "0%" FOR CONTRIBUTION


Placing Samples onto the Fluorimeter

  1. [Step One: A 160 microliter drop of water should be placed in between the first two rows of slides and the drop must be clear and round.]
  2. []
  3. [Instructions: Step three, in your own words]
  4. [Instructions: Step etc., in your own words]


Data Collection and Analysis

The images for the high, low, and zero Calf Thymus DNA were deleted and misplaced.


Calibrator Mean Values


calibration table


Calibration curves
Dot Plot 1 GCYLO.PNG Dot Plot 2 GCYLO.PNG

Positive PCR Control IMG 0652.jpg

Negative PCR Control IMG 0655.jpg


PCR Results: PCR concentrations solved

PCR table


PCR Results: Summary

527991 = 2653.632x+2542.374, x=198.011

  • Our positive control PCR result was 198.011 μg/mL
  • Our negative control PCR result was 55.592 μg/mL


Observed results

  • Patient 27800:

1-1: 177.01

1-2: 204.93

1-3: 174.42

Mean: 185.45

  • Patient 35993:

2-1: 41.308

2-2: 42.067

2-3: 55.749

Mean: 45.04


Conclusions

  • Patient 27800:

Patient 27800 has a mean concentration of 185.45 ug/mL, a mere 8 ug/mL away from the positive control, indicating that the patient contains the mutation indicated by the PCR test.

  • Patient 35993:

Patient 35993 has a mean concentration of 45.04, below the threshold for the negative control, indicating that the patient does not contain the mutation indicated by the PCR test.