BME103 s2013:T900 Group4 L2: Difference between revisions
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{| style="wikitable" width="700px" | {| style="wikitable" width="700px" | ||
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| [[Image:Group4Monkey2.jpg|120px|thumb|Name: Kinjal Ahir | | [[Image:Group4Monkey2.jpg|120px|thumb|Name: Kinjal Ahir <br> Procedure]] | ||
| [[Image: | | [[Image:133834815227.jpg|120px|thumb|Name:Zach Young <br>Initial Machine Testing]] | ||
| [[Image: | | [[Image:Superfunnypetmonkey01big.jpg|120px|thumb|Name: Anna Essex <br>Initial Machine Testing]] | ||
| [[Image: | | [[Image:Group4EasterBunny.jpg|120px|thumb|Name: Tuan Phan <br>Research & Design]] | ||
| [[Image:BME103_Group4Vergil_ODST.jpg| | | [[Image:BME103_Group4Vergil_ODST.jpg|120px|thumb|Name: Amelia Lax <br>Research and Design]] | ||
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'''SYBR Green Dye'''<br> | '''SYBR Green Dye'''<br> | ||
SYBR Green I is a fluorescent dye that binds to double stranded DNA | SYBR Green I is a fluorescent dye that binds to double-stranded DNA and fluoresces green when excited by blue light. SYBR green dye is very safe to users and cheap, making it ideal for basic DNA visualization. | ||
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'''Single-Drop Fluorimeter'''<br> | '''Single-Drop Fluorimeter'''<br> | ||
A fluorimeter is a device used to measure amounts of fluorescence. | A fluorimeter is a device used to measure amounts of fluorescence. Fluorimeters usually measure fixed wavelengths either with filters, monochromators, or fixed diodes. Fluorimeters have two light detectors; one detects absorbance while the other detects fluorescence emission. <br> | ||
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[[Image:Group4_Fluorometer.JPG]]<br> | [[Image:Group4_Fluorometer.JPG|500px]]<br> | ||
Drops of DNA solution on flourimeter slide | ''Drops of DNA solution on flourimeter slide'' | ||
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'''How the Fluorescence Technique Works'''<br> | '''How the Fluorescence Technique Works'''<br> | ||
The fluorescence technique measures the concentration of DNA in | The fluorescence technique measures the concentration of DNA in a solution by adding a fluorescent dye and measuring the amount of fluorescence. This is done using a single drop fluorometer with a Teflon-coated glass slide. The slide's surface is not completely covered; circles of bare glass remain, which water sticks to, allowing drops of liquid to stay in place. Drops of DNA solution are placed on the glass circles along with SYBR Green I. Blue LED light from the fluorometer is aimed at the drops, exciting the dye and causing it to fluoresce. This fluorescence is then analyzed to determine the concentration of DNA in the solution. | ||
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'''Calibration''' | '''Calibration''' | ||
* Place the smartphone on the cradle at a right angle | * Place the smartphone on the cradle at a right angle to both the table and the fluorimeter slide | ||
* | * Adjust the height of the fluorimeter slide if necessary so the camera is level with the drop | ||
* Distance between the smartphone cradle and drop = 4cm | * Distance between the smartphone cradle and drop = 4cm | ||
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{| {{table}} width=700 | {| {{table}} width=700 | ||
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| Calf Thymus DNA | | Calf Thymus DNA Solution Concentration (μg/mL) || Volume of the 2x DNA Solution (μL) || Volume of the SYBR GREEN I Dye Solution (μL) || Final DNA Concentration PicoGreen Assay (ng/mL) | ||
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| 5 || 80 || 80 || 2.5 | | 5 || 80 || 80 || 2.5 | ||
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'''Placing Samples onto the Fluorimeter''' | '''Placing Samples onto the Fluorimeter''' | ||
* Step one: | * Step one: Adjust camera settings and turn the flash off | ||
* Step two: | * Step two: Obtain calf thymus DNA, gloves, lab coat, glass slide, buffer tubes, and SYBR Green I | ||
* Step three: | * Step three: Insert Teflon-coated glass slide into fluorimeter | ||
* Step four: Drop 80μL of | * Step four: Drop 80μL of solution onto slide and position in front of blue light | ||
* Step five: Put | * Step five: Put dark box over fluorimeter and smartphone and take pictures of the drops | ||
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==Data Analysis== | ==Data Analysis== | ||
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[[Image:Negative DNA circle.jpg|500px]]<br> | [[Image:Negative DNA circle.jpg|500px]]<br> | ||
''Circle analysis of DNA negative drop (0μg/mL)'' | ''Circle analysis of DNA negative drop (0μg/mL)'' | ||
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[[Image:Positive DNA circle.jpg|500px]]<br> | [[Image:Positive DNA circle.jpg|500px]]<br> | ||
''Circle Analysis of DNA positive drop (5μg/mL)'' | ''Circle Analysis of DNA positive drop (5μg/mL)'' | ||
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As shown, the DNA positive drop is much brighter than the DNA negative drop. | |||
As shown, the DNA positive drop is much brighter. | |||
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'''Fitting a Straight Line'''<br> | |||
[[Image:LineFit.jpg|700px]] | |||
'' | ''General trendline of INTDENS readings vs. DNA concentration'' | ||
Latest revision as of 09:04, 2 April 2013
BME 103 Spring 2013 | Home People Lab Write-Up 1 Lab Write-Up 2 Lab Write-Up 3 Course Logistics For Instructors Photos Wiki Editing Help | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
OUR TEAM
LAB 2 WRITE-UPBackground InformationSYBR Green Dye Single-Drop Fluorimeter
How the Fluorescence Technique Works
ProcedureSmart Phone Camera Settings
Data AnalysisRepresentative Images of Samples
General trendline of INTDENS readings vs. DNA concentration
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