BME100 f2014:Group6 L5: Difference between revisions

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|- valign="top"
|- valign="top"
| [[Image:Prakriti_Picture.jpg|100px|thumb|Name: Prakriti Shukla <br>]]
| [[Image:Prakriti_Picture.jpg|100px|thumb|Name: Prakriti Shukla <br>]]
| [[Image:BME103student.jpg|100px|thumb|Name: student]]
| [[Image:GavinWhitePicture.jpg|100px|thumb|Name: Gavin White]]
| [[Image:BME103student.jpg|100px|thumb|Name: student]]
| [[Image:Galt Goettl.jpg|100px|thumb|Name: Galt Goettl]]
| [[Image:BME103student.jpg|100px|thumb|Name: student]]
| [[Image:IMG_4132.PNG|100px|thumb|Name: Laura Stokes]]
| [[Image:BME103student.jpg|100px|thumb|Name: student]]
| [[Image:CheetahPicture.jpeg|100px|thumb|Name: Paul Chua]]
| [[Image:BME103student.jpg|100px|thumb|Name: student]]
| [[Image:Redrosepicturefinal.jpeg|100px|thumb|Name: Yamilex Bustamante]]
|}
|}


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* Type of Smartphone: iPhone 5
* Type of Smartphone: iPhone 5
** Flash: Off
** Flash: Off
** ISO setting:
** ISO setting:n/a
** White Balance:  
** White Balance: n/a
** Exposure:
** Exposure:n/a
** Saturation:
** Saturation:n/a
** Contrast:
** Contrast:n/a




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<!-- INSTRUCTIONS: Type the distance between your phone cradle and the drop after the equal sign. -->
<!-- INSTRUCTIONS: Type the distance between your phone cradle and the drop after the equal sign. -->
* Distance between the smart phone cradle and drop =
* Distance between the smart phone cradle and drop = 4 cm




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{| {{table}} width=700
{| {{table}} width=700
|-
|-
| row 1 cell 1 || row 1 cell 2 || row 1 cell 3 || row 1 cell 4
| Concentration ofCalf Thymus DNA solution(micrograms/mL)|| Volume of theDNA solution (μL)|| Volume of the SYBR GREEN I Dye Solution (μL)|| Final DNA concentration in SYBR Green I solution (μg/mL)
|-
|-
| row 2 cell 1 || row 2 cell 2 || row 2 cell 3 || row 2 cell 4
| 5 || 80 || 80|| 2.5
|-
|-
| row 3 cell 1 || row 3 cell 2 || row 3 cell 3 || row 3 cell 4
| 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
 
|}
|}


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'''Placing Samples onto the Fluorimeter'''
'''Placing Samples onto the Fluorimeter'''
# ''[Instructions: Step one, in your OWN words]''
# "[Place the slide, rough side up, onto the fluorimeter]"
# ''[Instructions: Step two, in your own words]''
# "[Place 80 microliters of the SYBR Green Solution into the two middle circles of the slide]"
# ''[Instructions: Step three, in your own words]''
# "[Then place 80 microliters of the calibration solution into the same drop of the SYBR Green solution]"
# ''[Instructions: Step etc., in your own words]''
# "[Adjust the slide so the drop is lighted by the blue light of the fluorimeter]"


<br>
<br>
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<!-- INSTRUCTIONS: (1) Show ONE image where you drew a circle around the droplet with the freehand tool for any sample with *no* DNA. (2) Show ONE image where you drew a circle around the droplet with the freehand tool for a sample *with* DNA (positive signal). -If you include more than two images, you will not receive any additional credit. -->
<!-- INSTRUCTIONS: (1) Show ONE image where you drew a circle around the droplet with the freehand tool for any sample with *no* DNA. (2) Show ONE image where you drew a circle around the droplet with the freehand tool for a sample *with* DNA (positive signal). -If you include more than two images, you will not receive any additional credit. -->


'''Positive Sample'''
[[Image:Positivecontrolgroup6.png]]
'''Negative Sample'''
[[Image:Negativecontrolgroup6.png]]




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<!-- INSTRUCTIONS: Show a table for the ImageJ calf thymus DNA data. '''To save time on typing a new Wiki table from scratch''', use THIS TOOL to auto-generate a Wiki table: http://excel2wiki.net/wikipedia.php. Copy the headers and values from the Excel spreadsheet you made, paste them into the form field, click submit, copy the Wiki code that the tool generated, and replace TABLE GOES HERE (below) with your auto-generated code.  -->
<!-- INSTRUCTIONS: Show a table for the ImageJ calf thymus DNA data. '''To save time on typing a new Wiki table from scratch''', use THIS TOOL to auto-generate a Wiki table: http://excel2wiki.net/wikipedia.php. Copy the headers and values from the Excel spreadsheet you made, paste them into the form field, click submit, copy the Wiki code that the tool generated, and replace TABLE GOES HERE (below) with your auto-generated code.  -->


 
[[Image:PCRPRODUCT1.png]]
TABLE GOES HERE
[[Image:PCRPRODUCT2.png]]
 
[[Image:PCRPRODUCT3.png]]
[[Image:PCRPRODUCT4.png]]


'''Calibration curve'''<br>
'''Calibration curve'''<br>
<!-- INSTRUCTIONS: Place an image of your Excel plot with a line of best fit here. -->
<!-- INSTRUCTIONS: Place an image of your Excel plot with a line of best fit here. -->
 
[[Image:Concentrationgraphgroup6.png]]


'''PCR Results Summary'''
'''PCR Results Summary'''
<!-- INSTRUCTIONS: You completed 8 PCR reactions and used the SYBR Green I staining and imaging technique to measure the amount of amplified DNA in each PCR reaction. You used a standard curve (based on known concentrations of calf thymus DNA) to convert INTDEN values into DNA concentration. Your positive control and negative control samples should be used as '''threshold''' values for determining whether an unknown (patient) sample is truly positive or negative. Replace the underscore with your claculated initial concentration values.-->
<!-- INSTRUCTIONS: You completed 8 PCR reactions and used the SYBR Green I staining and imaging technique to measure the amount of amplified DNA in each PCR reaction. You used a standard curve (based on known concentrations of calf thymus DNA) to convert INTDEN values into DNA concentration. Your positive control and negative control samples should be used as '''threshold''' values for determining whether an unknown (patient) sample is truly positive or negative. Replace the underscore with your claculated initial concentration values.-->
* Our positive control PCR result was ____ μg/mL
* Our positive control PCR result was 5.44 μg/mL
* Our negative control PCR result was ____ μg/mL
* Our negative control PCR result was -3.19 μg/mL


<u>Observed results</u>
<u>Observed results</u>
<!-- INSTRUCTIONS: Replace the underscore with each patient ID. After the colon, write both a qualitative (what the images looked like) and a quantitative description (μg/mL) of what you observed -->
<!-- INSTRUCTIONS: Replace the underscore with each patient ID. After the colon, write both a qualitative (what the images looked like) and a quantitative description (μg/mL) of what you observed -->
* Patient _____ :  
* Patient 29148: The first sample was very green
* Patient _____ :
* Patient 35264 : The second sample had no color


<u>Conclusions</u>
<u>Conclusions</u>
<!-- INSTRUCTIONS: Compare each patient's results to the positive control value and the negative control value. Draw a final conclusion for each patient (positive or negative) and explain why you made that conclusion. -->
<!-- INSTRUCTIONS: Compare each patient's results to the positive control value and the negative control value. Draw a final conclusion for each patient (positive or negative) and explain why you made that conclusion. -->
* Patient _____ :
* Patient 29148: This patient was close to the positive control and would amplify nearly as much as the positive control.
* Patient _____ :
* Patient 35264 : This patient was close to the negative control and will not amplify as much




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<!-- INSTRUCTIONS: This content is from PCR Lab D. Write a summary, at least five sentences long, about the disease SNP in your own words. -->
<!-- INSTRUCTIONS: This content is from PCR Lab D. Write a summary, at least five sentences long, about the disease SNP in your own words. -->


SNP is a single nucleotide polymorphism. It occurs commonly within a population in which a single nucleotide (A, T, C, G) differs between members of a biological species. Almost all common SNPS occur within two alleles. SNPs occur within non-coding regions rather than coding regions. Genomic recombination and mutation rate can also vary SNP density. These genetic variations are exploited in DNA fingerprinting which is used in forensic science


'''Primer Design and Testing'''
'''Primer Design and Testing'''
<!-- INSTRUCTIONS: Write a short summary of the results of your primer test. Underneath your summary, include a screen capture of the results web page. You may crop the image so that it only includes the relevant information. -->
<!-- INSTRUCTIONS: Write a short summary of the results of your primer test. Underneath your summary, include a screen capture of the results web page. You may crop the image so that it only includes the relevant information. -->
 
The non-disease primer was successful while the disease primer did not have any results because of the SNPs in the disease primer.
 
[[Image:Nondiseaseprimerfinal.png]]
 
[[Image:Diseaseprimerfinal.png]]
 


<!-- Do not edit below this line -->
<!-- Do not edit below this line -->
|}
|}

Latest revision as of 21:09, 11 November 2014

BME 100 Fall 2014 Home
People
Lab Write-Up 1 | Lab Write-Up 2 | Lab Write-Up 3
Lab Write-Up 4 | Lab Write-Up 5 | Lab Write-Up 6
Course Logistics For Instructors
Photos
Wiki Editing Help


OUR TEAM

Name: Prakriti Shukla
Name: Gavin White
Name: Galt Goettl
Name: Laura Stokes
Name: Paul Chua
Name: Yamilex Bustamante


LAB 5 WRITE-UP

Procedure

Smart Phone Camera Settings

  • Type of Smartphone: iPhone 5
    • Flash: Off
    • ISO setting:n/a
    • White Balance: n/a
    • Exposure:n/a
    • Saturation:n/a
    • Contrast:n/a


Calibration

  • Distance between the smart phone cradle and drop = 4 cm


Solutions Used for Calibration

Concentration ofCalf Thymus DNA solution(micrograms/mL) Volume of theDNA 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 the slide, rough side up, onto the fluorimeter]"
  2. "[Place 80 microliters of the SYBR Green Solution into the two middle circles of the slide]"
  3. "[Then place 80 microliters of the calibration solution into the same drop of the SYBR Green solution]"
  4. "[Adjust the slide so the drop is lighted by the blue light of the fluorimeter]"


Data Analysis

Representative Images of Negative and Positive Samples

Positive Sample Negative Sample


Image J Values for All Calibrator Samples

Calibration curve

PCR Results Summary

  • Our positive control PCR result was 5.44 μg/mL
  • Our negative control PCR result was -3.19 μg/mL

Observed results

  • Patient 29148: The first sample was very green
  • Patient 35264 : The second sample had no color

Conclusions

  • Patient 29148: This patient was close to the positive control and would amplify nearly as much as the positive control.
  • Patient 35264 : This patient was close to the negative control and will not amplify as much




SNP Information & Primer Design

Background: About the Disease SNP

SNP is a single nucleotide polymorphism. It occurs commonly within a population in which a single nucleotide (A, T, C, G) differs between members of a biological species. Almost all common SNPS occur within two alleles. SNPs occur within non-coding regions rather than coding regions. Genomic recombination and mutation rate can also vary SNP density. These genetic variations are exploited in DNA fingerprinting which is used in forensic science

Primer Design and Testing The non-disease primer was successful while the disease primer did not have any results because of the SNPs in the disease primer.

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