BME100 s2018:Group3 W1030 L5

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OUR TEAM

Name: Alicia Salas
Name: Anastasia Hancock
Name: John Le
Name: Mikayla Gardes


LAB 5 WRITE-UP

PCR Reaction Report

Our experience with pipetting was mixed all around. One was experienced with pipettes since they volunteered at a local clinic, another was new to the concept so it was rather difficult at first trying to remember the order of pipetting. The rest of us had been introduced it in a lab from another class. The pre-lab was useful because we were able to refresh on the idea and how it was used. It is easy to get mixed up and we did not want that because it could potentially ruin the samples. The difference between the two stops was what step you were on obtaining the samples. The first stop is to collect the samples at the preferred setting and if one were to go all the to the second stop while collecting the sample, then the sample would come back into the pipette rather the tip. The second stop is used while delivering samples by completely expelling everything in the tip. The final reaction did have the same amount of liquid at first but after close observation, there was a drop or two left in the sample tubes. Although the mircopipette is extremely accurate, it is known on not getting every single drop. The labeling scheme was kept the same to avoid confusion throughout the lab.

Fluorimeter Procedure

Imaging set-up

  1. While using a pipettor, take 160 microliters of water and place it in the middle of the first two rows. The drop must round and pinned.
  2. Flip the switch which allows the Blue LED and the excitation light.
  3. Camera is powered on and is adjusted accordingly.
  4. Adjust the Fluorimeter accordingly where the camera can visibly see the drop straight on rather at angle.
  5. Adjust the camera where it can zoom in as much as possible without becoming blurry. Best distance would 4 cm away.
  6. Record distance from camera and drop.

Placing Samples onto the Fluorimeter

  1. Set the micropipettor to 120 microlitters.
  2. The disposable tip is attached to the pipette.
  3. Collect 100 microlitters of PCR into the Buffer tubes given.
  4. Dispose the Mircopipette tip.
  5. Make sure the DNA is thoroughly mized with the Buffer.
  6. Repeat steps 3 though 5 until all DNA samples are used.



Data Collection and Analysis

Images of High, Low, and Zero Calf Thymus DNA


Group3 W1030 5-1.png High 5-1 Sample Group3 1030 0.5-1.PNG Low 0.5-1 Sample H2O33.png Zero H2O Sample


Calibrator Mean Values


Calibrator Mean Values.PNG

Calibration curves

Calibration curve 1 Group3.PNG Calibration Curve 2 Corrected.PNG



Images of Our PCR Negative and Positive Controls

Positive1.PNG Positive Neg-12.PNG Negative


PCR Results: PCR concentrations solved PCR Solved Corrected.PNG


PCR Results: Summary

  • Our positive control PCR result was 47.196 μg/mL
  • Our negative control PCR result was 0.93924 μg/mL


Observed results

  • Patient 74852 : The patient has a green color when placed through the laser. 80 mL of the patient was placed in the sample.
  • Patient 42241 : The patient has a dark blue color when placed through the laser. 80 mL of the patient was placed in the sample.


Conclusions

  • Patient 74852 :Positive, there is a green glow when exposed to the light.
  • Patient 42241 :Negative, there is no glow present when exposed to the light.