BME100 s2015:Group2 9amL5

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Contents

OUR TEAM

Ryan BridgesRole(s)
Ryan Bridges
Role(s)
Name: Michael BejaranoRole(s)
Name: Michael Bejarano
Role(s)
Name: Virginia FernandezRole(s)
Name: Virginia Fernandez
Role(s)
Morgan JohnsonRole(s)
Morgan Johnson
Role(s)


LAB 5 WRITE-UP

Procedure

Smart Phone Camera Settings

  • Type of Smartphone: Samsung Galaxy S5
    • Flash: Off
    • ISO setting: 2000
    • White Balance: Auto
    • Exposure: 0.0
    • Saturation: N/A
    • Contrast: Auto


Calibration

  • Distance between the smart phone cradle and drop = 5cm


Solutions Used for Calibration

Initial Concentration of 2X Calf Thymus DNA solution (micrograms/mL) Volume of the 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. Place a 80 microliter drop of SYBR Green I on the front center of the slide.
  2. Add a 80 microliter drop of sample DNA to the SYBR Green I drop.
  3. Ensure no light enters the box and the LED shines directly on the drop.
  4. Center camera cradle and focus camera clearly on the drop.
  5. Set timer and close box.
  6. After picture three pictures have been taken, remove box and remove sample with micropipettor.
  7. Repeat steps for all samples.



Data Analysis

Representative Images of Negative and Positive Samples

Negative Image:Capture9.jpg

Positive Image:Capture8.jpg


Image J Values for All Calibrator Samples


Image:Capture_sample_table.jpg

Image:LAB_5EXC.bmp

Image:LAB_5CAL.bmp

Image:LAB_5DB.bmp


Calibration curve

Image:LAB_5CAC.bmp


PCR Results Summary

  • Our positive control PCR result was 0.12189 μg/mL
  • Our negative control PCR result was 0.065342 μg/mL

Observed results

  • Patient 15062 : The image had a small amount of green, the concentration was found to be 0.13133 μg/mL
  • Patient 95748 : The image has no green. The concentration was found to be 0.03870 μg/mL

Conclusions

  • Patient 15062 : Based on the concentration of DNA in the sample compared to the positive control value and the negative control value, patient 15062 was positive.
  • Patient 95748 : Based on the concentration of DNA in the sample compared to the positive control value and the negative control value, patient 95748 was negative.




SNP Information & Primer Design

Background: About the Disease SNP

A nucleotide is a bundle of amino acids that serve as building blocks for DNA. A polymorphism is a variation of phenotypes within a select group population. The single nucleotide polymorphism (SNP) rs268 is found in Homo sapiens (Human). The chromosome this variation is located on is 8. The listed clinical significance of this SNP is pathogenic. The genes this SNP is associated with are NHGRI, GWAS, and PheGenl. Coronary heart disease is linked to this SNP. LPL stands for lipoprotein lipase. The functions of LPL include: apolipoprotein binding, heparin binding, and lipoprotein lipase activity. An allele is a variation of a gene usually brought to pass by a mutation. The sequence of the disease-associated allele is AGT. The position of the SNP is 19956018.


Primer Design and Testing

The results of the primer test of the DNA sequences of diseased and non-diseased primers were successful in that the expected outcome was achieved. A match was found with the non-diseased primer because the human genome produces this healthy primer on its own while no match was found when the diseased primer was tested because the human genome produce the mutation that was present in the diseased primer.

Image:LAB_5M.bmp Image:LAB_5NM.bmp

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