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
[Instructions: Step one, in your OWN words]
[Instructions: Step two, in your own words]
[Instructions: Step three, in your own words]
[Instructions: Step etc., in your own words]
Data Collection and Analysis
Images of High, Low, and Zero Calf Thymus DNA
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
Images of Our PCR Negative and Positive Controls
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.
BME 100 Fall 2015
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