BME103 s2013:T900 Group4 L2: Difference between revisions
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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 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 | 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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{| {{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) | ||
|- | |- | ||
| 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: Obtain calf thymus DNA, gloves, lab coat, glass slide, buffer tubes, and SYBR Green I | * 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 solution onto slide in front of blue light | * Step four: Drop 80μL of solution onto slide and position in front of blue light | ||
* Step five: Put dark box over fluorimeter and smartphone and take pictures of the drops | * Step five: Put dark box over fluorimeter and smartphone and take pictures of the drops | ||
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As shown, the DNA positive drop is much brighter. | As shown, the DNA positive drop is much brighter than the DNA negative drop. | ||
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'''Fitting a Straight Line'''<br> | |||
[[Image:LineFit.jpg|700px]] | |||
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''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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