Download limma: Linear Models for Microarray Data User's Guide

Chapter 4
Reading Two-Color Data
4.1
Scope of this Chapter
This chapter is for two-color arrays. If you are using Affymetrix arrays, you should use the affy
or affyPLM packages to read and normalize the data. If you have single channel arrays other
than Affymetrix, you will need to the read the intensity data into your R session yourself using
the basic R read functions such as read.table. You will need to create a matrix containing
the log-intensities with rows for probes and columns for arrays.
4.2
Recommended Files
We assume that an experiment has been conducted with one or more microarrays, all printed
with the same library of probes. Each array has been scanned to produce a TIFF image. The
TIFF images have then been processed using an image analysis program such a ArrayVision,
ImaGene, GenePix, QuantArray or SPOT to acquire the red and green foreground and background intensities for each spot. The spot intensities have then been exported from the image
analysis program into a series of text files. There should be one file for each array or, in the
case of Imagene, two files for each array.
You will need to have the image analysis output files. In most cases these files will include
the IDs and names of the probes and possibly other annotation information. A few image
analysis programs, for example SPOT, do not write the probe IDs into the output files. In
this case you will also need a genelist file which describes the probes. It most cases it is also
desirable to have a targets file which describes which RNA sample was hybridized to each
channel of each array. A further optional file is the spot types file which identifies special
probes such as control spots.
4.3
The Targets Frame
The first step in preparing data for input into limma is usually to create a targets file which
lists the RNA target hybridized to each channel of each array. It is normally in tab-delimited
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