Table 1.
Transcript-specificity of HG-U133A probes to Ensembl transcripts.
Figure 1.
Intensity histograms of several probe categories.
Gene-specific and transcript-specific probes show more high intensity signals as compared to non-perfect or no-match probes. a) Probe intensities of one microarray (LD2) were RMA-background-adjusted and logarithmized (base 2). Probes were assigned to categories shown in the legend as defined in the results section. The normalized intensities of each category are put into 100 evenly-spaced intervals and plotted as histogram. The probe intensity 95%-quantile of Affymetrix-defined negative control probe sets is plotted in yellow. b) As the number of probes varies between the different categories it is hard to compare the shapes of the distributions in a). Therefore, the histograms were normalized with their number of probes in this plot. Averaging over multiple arrays instead of analyzing one array gives similar results.
Table 2.
Genes having transcripts with transcript-specific probes.
Figure 2.
Comparison of alternative transcript abundance in microarray and real-time RT-PCR.
On the left microarray signal intensities are shown for the genes PPM1A and PLG. Confirmatory real-time RT-PCR data are shown on the right. Lowly and highly abundant transcripts of the genes PPM1A and PLG were measured using microarrays in LD tissue. Transcript-specific probe intensities were background-adjusted and quantile-normalized using RMA. These are shown as single probe values (triangles) and, furthermore, as summarized transcript-specific probe set values (dots). In addition, the transcripts were quantified in the unaffected part of TN tissue using real-time RT-PCR. Real-time RT-PCR data are normalized to the transcript with lower abundance.
Figure 3.
Microarray and real-time RT-PCR transcript measurements of PLG and ABCA1 in two patient cohorts.
For the gene PLG, transcript PLMN_HUMAN was repressed in DD compared to LD controls, while Q5TEH_HUMAN showed no differential expression. Real-time RT-PCR measurements on the same tissues were in agreement with these findings. For gene ABCA1, transcript ABCA1_HUMAN was induced in DN compared to LD, and Q5VYS0_HUMAN was not regulated. Real-time RT-PCR measurement confirmed the induction of ABCA1_HUMAN. Q5VYS0_HUMAN was expressed at a too low level to be measured (value 0). Real-time data are normalized to the cohort with lower abundance.
Table 3.
Significance analysis of intensity differences between alternative transcripts of two genes.
Table 4.
Primers used for real-time RT-PCR.