BME100 s2015:Group5 9amL5

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Contents

OUR TEAM

Name: Jacob Licko
Name: Jacob Licko
Name: Wei Lu
Name: Wei Lu
Name: Daniela Romero
Name: Daniela Romero
Name: Khalil Khoury
Name: Khalil Khoury
Name: Stephen Belikoff
Name: Stephen Belikoff


LAB 5 WRITE-UP

Procedure

Smart Phone Camera Settings

  • Type of Smartphone: iPhone6
    • Flash:Automatic
    • ISO setting:Normal
    • White Balance: 0 (neutral)
    • Exposure:0 (neutral)
    • Saturation:0 (neutral)
    • Contrast: 0 (neutral)


Calibration

  • Distance between the smart phone cradle and drop = 3 inches


Solutions Used for Calibration

Initial Concentration of 2X Calf Thymus DNA Solution (micrograms/mL) Volume ofthe 2X DNA Solution (μL) Volume of the SYBR GREEN I Dye Solution (μL) Final DNA concentration in SYBR Green I Solution μg/mL
5 80 80 2.5
2 80 80 1
1 80 80 0.5
0.5 80 80 0.25
0.25 80 80 0.125
0 80 80 0



Placing Samples onto the Fluorimeter

  1. Dispense 80μL of SYBR GREEN onto the first two middle dots of the glass tray, make sure the rough end is pointed up.
  2. Dispense 80μL of the Dalf Thymus DNA solution onto the SYBR GREEN drop.
  3. Close the system that the tray is on so that little to no light can penetrate the drop.
  4. Turn on the light and take three bursts of pictures of the drop


Data Analysis

Representative Images of Negative and Positive Samples

Negative:

Positive:

Image J Values for All Calibrator Samples


Final DNA concentration in SYBR Green I solution (µg/mL) Area Mean Pixel Value RAWINTDEN of the drop RAWINTDEN of the background RAWINTDEN drop - background
Water31172167.519522189427414782480416
Water₂3989657.97223128681544132158455
.25 Concentration36784200.67373815532922177089336
.25 Concentration₂39608102.80740719881689193903069
.5 Concentration31944208.35466556502603656395285
.5 Concentration₂35124112.85339638431428643820979
1 Concentration36500209.04876302472965257333722
1 Concentration₂39588128.89751027931633404939453
2 Concentration37336236.59188333593298388503521
2 Concentration₂40188194.85278307071749877655720
5 Concentration36200234.43684865852913528195233
5 Concentration₂31680219.34869489381531546795784


Final DNA concentration in SYBR Green I soluction (µg/mL RAWINTDEN drop - background ' ' Standard Deviation
12Mean
Patient 1-1653093628128534671894.52629081.702
Patient 1-2543890027176594078279.51924207.964
Patient 1-36379485360267949910821963498.353
Patient 2-16958685285764949081672899870.365
Patient 2-2397645188920986434274.53475887.328
Patient 2-3912890856707777399842.52445267.88


Calibration curve
Image:Fjdh.PNG


PCR Results Summary

  • Our positive control PCR result was 80 μg/mL
  • Our negative control PCR result was 80 μg/mL

Observed results

  • Patient 59797 : The solution was clear, or had a hue of blue due to the flourescent light. This was observed in the three duplicates of the first patient's DNA.
  • Patient 79049: The solution was clear, or had a hue of blue due to the flourescent light on 2 of the three duplicates. However for the 2nd dupliate, 2-2, the drop was a green color.

Conclusions

  • Patient 59797 : The patient tested negative for diabetes.
  • Patient 79049 : The patient testes positive for diabetes.




SNP Information & Primer Design

Background: About the Disease SNP The SNP was a Pathogenic disease that has an association with coronary heart disease. The variation is found in humans and affects the NHGRI, GWAS, and PheGenl genes.


Primer Design and Testing

For the non-disease DNA code, the result was matched 220bp because there was no mutation on the 3 prime end of the code. For the disease DNA code, the result was 'no matches' because there was a mutation at the 3 prime end where the A nucleotide aligned to a G nucleotide. Thus causing the mutation and the 'no matches'.

Non-Disease: Image:Non-Diseas.jpg

Disease: Image:Disease.jpg



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