Dahlquist:GenePix Pro Software Protocol

From OpenWetWare

(Difference between revisions)
Jump to: navigation, search
(instructions for downloading files)
(start instructions for TOD6)
Line 64: Line 64:
* Name the file: Chip #_Year-Month-Date  
* Name the file: Chip #_Year-Month-Date  
* The PDF file will either open on its own or you need to open it from the file you saved it to
* The PDF file will either open on its own or you need to open it from the file you saved it to
 +
 +
== Find the spot corresponding to TOD6 on the microarray ==
 +
 +
* Go to [http://www.yeastgenome.org/cgi-bin/locus.fpl?locus=TOD6 TOD6 page at the ''Saccharomyces'' Gene Database]

Revision as of 12:41, 14 April 2010

Contents

Using GenePix Pro

Biology 478/Spring 2010 students:

  • Follow this link to access your microarray image and .gal files from LionShare: https://lionshare.lmu.edu/Users/kdahlqui/BIOL478
  • Download both the 532 and 635 wavelength files for your group onto your Desktop
    • HINT: Right-click on the link and select the menu item "Save target as" or "Save link as" to download the file. If you just click on the link, your browser will attempt to open it without downloading
  • Red, orange, yellow, green, and pink groups, download the file "yeast_GCAT_Fall2006_gal.gal" to your Desktop
  • Blue group, download the file "GCAT_384_illumina_Spring2007.gal" to your Desktop

Gridding and Generating Intensity Data

  • Launch GenPix Pro (select Analysis Only)
  • On the right hand side of the screen, select the File icon
    • Select File > Open Images
    • Navigate to the folder containing your .tif images
    • Hold down the Control key while clicking to select both the 532 and 635 wavelength files
    • Click on the Open button
  • Use the brightness and contrast sliders on the upper left hand side to adjust your image to see all of the spots
  • You can use the magnifying glass icon to zoom in on an area of the image, and the magnifying glass return icon to go back to the whole chip image
    • Zoom in so that you can see the entire top block of grids on your screen
  • On the right hand side of the screen, select the File icon
    • Select Load Array List
    • Red, orange, yellow, green, and pink groups, use the file "yeast_GCAT_Fall2006_gal.gal"
    • Blue group, use the file "GCAT_384_illumina_Spring2007.gal"
    • Click the Open button
  • Using the icon that has an arrow and a square, drag the grids so that they are approximately aligned with the top block of spots
    • Click on the icon that looks like a compass, or a circle with cross-hairs, and select Find Array
    • Click on the icon again, and select Find All Blocks
    • Click on the icon again, and select Align Features in All Blocks
  • On the right hand side of the screen, select the icon that looks like a table of data (it says B, C, R at the top of it)
  • In the Results tab, click on the button to Configure Normalization
    • Make sure that "Ratio-based" is selected and it says that the "Ratio of medians" of "all of the features" is equal to "1"
    • Click the Normalize button
  • Click on the File icon on the right hand side and select, Save Results As, and save your results.
  • Repeat this procedure with the bottom block on the slide.

Generating an Array Quality Report

  • Launch GenePix Pro (select Analysis Only)
  • On the right hand side of the screen, select the File icon
  • File > Open Results
  • Select the desired file
  • Select Report tab at the top of the screen
  • On the left side of the screen, under Navigate, select the back arrow or home icon
  • Select Array Quality Control
  • Adjust Vital Statistic Thresholds values to:
Quality Control Limits
Median signal to background > 2.5
Mean of Median background < 500
Median Signal to Noise	> 4
Median % >B + 1 StdDev	> 90
Feature variation < 0.5
Background Variation < 1.2
Features with Saturated pixels	< 3.3%
Not Found < 18%
Bad Features < 7%
  • Select Start
  • Select Show Printable Version
  • File > Print
  • Under Select Printer > Select Adobe PDF
  • Select Print
  • Save on the Desktop
  • Name the file: Chip #_Year-Month-Date
  • The PDF file will either open on its own or you need to open it from the file you saved it to

Find the spot corresponding to TOD6 on the microarray

Personal tools