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Fig. 2. Replicate hybridizations and reciprocal dye labeling experiments. (A)
Duplicate hybridizations were performed for each time point in the first two
temperature cycles. Expression data were filtered by accepting only spots that
changed over twofold in at least 1 time point. These data are not corrected
relative to t=0, but they are median centered. Cy3 was used to label
the reference sample while Cy5 was used to label the experimental sample in
the forward (F) hybridizations. The dyes were reversed (R) in the second set
of hybridizations. Visual inspection reveals a strong relationship between the
two sets of hybridizations, especially for spots that change greater than
twofold compared to the reference. (B) There is a strong correlation between
the data for the forward and reverse hybridizations (r=0.96) for
spots on the array that change more than twofold (N=831). The
equation for the regression line on the graph is
y=1.061x+0.04056. These data indicate that there is no
significant dye bias in this data set. Further, it supports the conclusion
that changes in gene expression observed in this study are not likely to be
due to variation in hybridizations, but instead are biologically relevant.
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