BME100 f2015:Group16 1030amL5

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OUR TEAM

Steven Rojo
Jacob Baca
Javon Boaz
Sudarshan Kandel
Aditya Konduri
Nicolas March


LAB 5 WRITE-UP

PCR Reaction Report

Group 16 had a slight trouble with the lab, due to an over abundance of reagent being added to the negative. This occurrence slightly modified our end results of this one specific test, but otherwise one of the teammates who had experience in a laboratory, was able to proceed with ease. This was further supported by the prelab materials, that laid out the plan for the lab in an easy-to-follow guide. The process of have the solution go up the pipette was found to be easy, despite the temperamental aspect of a first and second release, due to the user being experience in a proper laboratory environment.

Fluorimeter Procedure

Smart Phone Camera Settings

  • Type of Smartphone: Samsung Galaxy S5
    • Flash: Inactivated
    • ISO setting: 800
    • White Balance: Auto
    • Exposure: Default
    • Saturation: Default
    • Contrast: Default


Camera set-up
We used a stand to hold the smartphone steady, and used the timer feature to take the picture. We found the distance between the camera and fluorimeter most suitable for a focused photo. When replacing the sample we lifted the box to avoid changing the position of the flourimeter and camera.

  • Distance between the smart phone cradle and drop = 16.5 cm


Placing Samples onto the Fluorimeter

  1. Utilizing a micropipette, draw a sample of SYBR Green dye, measuring 80 mL, and inject the dye into the center of the slide, thus forming a drop.
  2. Utilizing a micropipette, draw a sample of the calf thymus sample solutions, again measuring 80 mL. Follow this up by delicately ejecting the solution onto the previous made drop, forming one single large drop.
  3. Align the slide such that the blue light travels through the center of the dot and is focused onto the black fitting on the other side.
  4. After the pictures needed are taken, use a provided pipette to remove the "drop" of solution from the slide.


Data Collection and Analysis

Images of High, Low, and Zero Calf Thymus DNA

High Concentration (5 μg/mL sample)

Viper.jpg

Low Concentration (.5 μg/mL sample)

Hulk1.jpg

Zero Concentration (0 μg/mL sample)

Meagaman.jpg

Calibrator Mean Values


Straightottacompton.jpg


Calibration curves
Dot 1.jpg Dot 2.jpg


Images of Our PCR Negative and Positive Controls

Negative Control
Positive Control


PCR Results: PCR concentrations solved

Fme.jpg


PCR Results: Summary

  • Our positive control PCR result was 42.05 μg/mL
  • Our negative control PCR result was -.7824 μg/mL


Observed results

  • Patient 17821: The droplet is an oblong blue shape with streaks of dark blue running vertical with the droplet. There also exists a black disc in the center running horizontal with the droplet.
  • Patient 75677: The droplet is an oblong blue shape with a black stripe running horizontal with the droplet.


Conclusions

  • Patient 17821: This patient's results for the concentration of DNA was found to be less than the initial concentration, thus resulting in a negative result which would not allow for the single nucleotide to show polymorphism.
  • Patient 75677: This patient's results for the concentration of DNA was found to be less than the initial concentration, thus resulting in a negative result which would not allow for the single nucleotide to show polymorphism.