BME100 f2015:Group14 1030amL5

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OUR TEAM

Name: Raymond E. Rivera
Name: Ethan B. Marschall
Name: Laura A. Roa
Name: Raul J. Ramirez
Name: Ian M. Wale
Name: Lauren T. Howard


LAB 5 WRITE-UP

PCR Reaction Report

Our team was able to use the pipette successfully and had no trouble with it during part C of our lab. The pre-lab video did help us due to the fact that explained how to use it thoroughly and each one of us knew how to do so. The first stop on the pipette allows you to retrieve liquid or any substance you are trying to reach inside of the pipette. The second stop is when you are releasing the material in the pipette into a tube, cylinder, etc. Our team understood this from the pre-lab reading and had no trouble understanding it. The final reactions did have the same amount of liquid. After using the pipette to combine the DNA samples and PCR reaction mix, each tube we had labeled had the same amount of liquid. In addition, there was no liquid left in either of the tubes that contained DNA samples and PCR reaction mix. The labeling scheme we used was provided in the lab workbook; we used the same names the workbook had to label our samples so that we did not have to change our labeling scheme.

Fluorimeter Procedure

Smart Phone Camera Settings

  • Type of Smartphone: Iphone 5S
    • Flash: not on
    • ISO setting: automatic
    • White Balance: automatic
    • Exposure: automatic
    • Saturation: automatic
    • Contrast: automatic


Camera set-up

Place the camera in the cradle at a right angle to the slide. Make sure the camera views the droplet from the side and is at least 4 centimeters away from the droplet.

  • Distance between the smart phone cradle and drop =

7 cm between the phone cradle and drop.

Placing Samples onto the Fluorimeter

  1. [Instructions: Step one, in your OWN words]
  2. [Instructions: Step two, in your own words]
  3. [Instructions: Step three, in your own words]
  4. [Instructions: Step etc., in your own words]


Data Collection and Analysis

Images of High, Low, and Zero Calf Thymus DNA

Gr14Capture C1 1867.PNG Capture C4 1856.PNG Capture C6 1850.PNG

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 ' ' MEAN Standard Deviation
Image 1 Image 2 Image 3
5 2.5 C-1 12470521 12512906 12494853 12492760 21269.87407
2 1 C-2 8726435 8777960 7128161 8210852 937991.7684
1 0.5 C-3 7439024 7569181 5568458 6858887.667 1119438.142
0.5 0.25 C-4 6476560 6485218 4850071 5937283 941563.163
0.25 0.125 C-5 4815437 4806830 3160825 4261030.667 952815.7754
0 0 C-6 3363487 3104174 2441282 2969647.667 475592.8051


Calibration curves

DotPlot1.PNG DotPlot2.PNG

Images of Our PCR Negative and Positive Controls

Capture negative 1870.PNG Capture positive 1872.PNG

PCR Results: PCR concentrations solved

PCR Product TUBE LABEL MEAN (of RAWINTDEN DROP - BACKGROUND) PCR Product Concentration (µg /mL) Total Dilution Initial PCR Product Concentration (µg /mL)
G14 + 9608646.333 2.36382305 12 28.3658766
G14 - 4761881 0.389167145 12 4.670005736
G14 1-1 3856448 0.020278103 12 0.243337233
G14 1-2 3698200.667 -0.044194594 12 -0.530335127
G14 1-3 3296592.667 -0.207816629 12 -2.493799551
G14 2-1 3259902.333 -0.222764905 12 -2.673178857
G14 2-2 3062879.667 -0.303035342 12 -3.636424103
G14 2-3 2904924.333 -0.367389073 12 -4.408668873


PCR Results: Summary

  • Our positive control PCR result was 28.37 μg/mL
  • Our negative control PCR result was 4.67 μg/mL


Observed results

  • Patient 69477 : The image is a circular object with light white shade but more dark unlike the first one. The observed quantitative data was 28.37 ug/ml.
  • Patient 37835 : The image is a ciruclar object that is shaded very dark and has a quantitative number of 4.67 ug/ml.


Conclusions

  • Patient 69477 : Negative. The initial values came out negative, so no PCR product was there.
  • Patient 37835 : Negative. The initial values came out negative, so no PCR product was there.