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Suppose there is an experiment designed to detect transcripts showing differential expression in two tissue groups: liver vs muscle. The alt-splicing ANOVA dialog allows you to specify the ANOVA model that is tissue, the alt-splicing factor is chosen from the ANOVA factor(s), so the alt-splicing factor is also tissue (Figure 1). Click Next to setup the comparisons (Figure 2)

 

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SubtitleText Alt-splicing ANOVA comparisons setup dialog AnchorName Alt-splicing-contrast

The subgroups of the alt-splicing factor will be displayed in the left panel; click to select a subgroup name and move it to one of the panels on the right. The fold change calculation on the comparison will use the group in the top panel as numerator, and the group in the bottom panel as denominator. Click on Add comparison button to add on comparison to the comparisons table.

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Alt-splicing ANOVA report

This this analysis, only genes with more than one transcripts will be included in the calculation. The report format is the same as ANOVA report, each row represent a transcript, besides statistics information on the specified comparisons, there is also Alt-splicing information at the right end of the table, this information is represented by the p-value of interaction of transcript ID with alt-splicing factor, transcripts of the same gene should have the same p-value. Small p-value indicate significance of the gene undergo alt-splicing, by default the table is sorted based on this p-value (Figure 4).

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SubtitleText Alt-splicing report. Clicking on the column header sorts the table. Panel on the left filters the table AnchorName Alt-splicing report
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In the above screenshot example, the alt-splicing p-value of gene SLC25A3 is very small which indicating this gene showing alt-splicing based on the tissue. There are 3 splicing variants in this gene: NM_213611, NM_005888 and NM_002635.  NM_005888 has much higher expression in muscle comparing to liver, but NM_002635 has much lower expression in muscle comparing to liver, the fold changes are opposite. When you click on the Browse to location view (Image Added) to visualize the difference (Figure 5).

 

 

 

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SubtitleText Use isoform proportion track in chromosome view to visualize the alt-splicing gene AnchorName Alt-splicing-chrview
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The 3rd exon is different between NM_005888 and NM_002635. Muscle primarily expressed in NM_005888 while liver primarily expressed in NM_002635.

 

 

Additional assistance
https://documentation.partek.com/

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