BME100 s2015:Group5 9amL5

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OUR TEAM

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

Negativegroup5.JPG

Positive:

Positivegroup5.jpg

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
Water 31172 167.519 5221894 2741478 2480416
Water₂ 39896 57.972 2312868 154413 2158455
.25 Concentration 36784 200.673 7381553 292217 7089336
.25 Concentration₂ 39608 102.807 4071988 168919 3903069
.5 Concentration 31944 208.354 6655650 260365 6395285
.5 Concentration₂ 35124 112.853 3963843 142864 3820979
1 Concentration 36500 209.048 7630247 296525 7333722
1 Concentration₂ 39588 128.897 5102793 163340 4939453
2 Concentration 37336 236.591 8833359 329838 8503521
2 Concentration₂ 40188 194.852 7830707 174987 7655720
5 Concentration 36200 234.436 8486585 291352 8195233
5 Concentration₂ 31680 219.348 6948938 153154 6795784


Final DNA concentration in SYBR Green I soluction (µg/mL RAWINTDEN drop - background ' ' Standard Deviation
1 2 Mean
Patient 1-1 6530936 2812853 4671894.5 2629081.702
Patient 1-2 5438900 2717659 4078279.5 1924207.964
Patient 1-3 6379485 3602679 4991082 1963498.353
Patient 2-1 6958685 2857649 4908167 2899870.365
Patient 2-2 3976451 8892098 6434274.5 3475887.328
Patient 2-3 9128908 5670777 7399842.5 2445267.88


Calibration curve
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: Non-Diseas.jpg

Disease: Disease.jpg