BME100 f2013:W900 Group10 L5
BME 100 Fall 2013 | 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
LAB 5 WRITE-UPBackground InformationSYBR Green Dye SBYR green dye is a cyanine dye used as a nucleic acid stain. When bound to DNA it absorbs blue light and emits green light. The stain binds to double stranded DNA at very high levels and binds to single stranded DNA at much lower levels. This allows us to measure the amount of double stranded DNA while getting minimal noise on our signal from the presence of single stranded DNA. It can also stain RNA at lower levels, but this is not important for our experiment (in which we know our sample does not contain RNA). It is also the most sensitive stain available for detecting double stranded DNA during PCR.
This device is used to excite the stain molecules in order to generate the signal (green light) for us to capture. The single-drop fluorimeter is designed to hold a single drop of the sample and pass the wavelength (~497nm) of light required to excite the stain molecules through the sample. [Instructions: A description of the single-drop fluorimeter device. Add a PHOTO for bonus points]
A droplet is placed on the hydrophobic slide which allows the droplet to hold its spherical shape. The sample droplet is then exposed to an ultraviolet light light beam to excite the stain molecules which proceed to emit green light (our signal). We capture this signal (green light) with our smartphone camera. In theory, the amount of signal captured by our camera should be proportional to the concentration of DNA in the sample (the pictures must be filtered so that only the amount of green light captured is taken into consideration). Therefore, once we have calibrated our camera with known concentrations of DNA we should be able to compare signal strengths (green light emittance) from unknown DNA concentration samples with our calibration data to accurately estimate the DNA concentration in that sample. [Instructions: In your own words]
ProcedureSmart Phone Camera Settings
Calibration [Instructions: Describe how to set up your camera in front of the fluorimeter. Add a PHOTO of this set-up for bonus points.] The camera was placed upright in the supplied stand. The camera lens was positioned 6cm from the droplet in order to get the closest view possible while still being within the focus range of the camera. The height of the fluorimeter was adjusted so that the lens of the camera was at the same height as the droplet to ensure a full side view.
Solutions Used for Calibration
Placing Samples onto the Fluorimeter
Data AnalysisRepresentative Images of Samples The picture below shows the control case where no DNA is present in the sample The picture below shows a test case where DNA is present in the sample
Image J Values for All Samples [Instructions: See worksheet page 8. To save time on typing a new Wiki table from scratch, use THIS TOOL to auto-generate a Wiki table: Excel-to-Wiki Converter. 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.]
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