Moderator: Benn
Benn ♦ 7.8k
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... How did you do enrichment, with which package? It might be that your BLAST2GO annotation is older than the GO database that was used for enrichment analysis. The GO database is very dynamic and changes with every update. So please explain more about how you did the enrichment analysis. ...
written 11 days ago by
Benn ♦ 7.8k
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... Two-channel (color) microarrays are usually normalized with LOESS, but one-channel arrays usually only with a between-array method (like quantile). The best tool to use is [limma][1], you can read in the manual how to normalize and analyze these different array types, including case studies with scr ...
written 11 days ago by
Benn ♦ 7.8k
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... To do GO enrichment analysis for more groups in parallel you can try [clusterProfiler][1]. I used it once to do GO enrichment analysis for 6-7 groups simultaneously.
[1]: https://bioconductor.org/packages/release/bioc/vignettes/clusterProfiler/inst/doc/clusterProfiler.html ...
written 12 days ago by
Benn ♦ 7.8k
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Comment:
C: How to normalize my rna seq data?
... You could use [voom][1] normalization from limma, and add the quantile normalization in there with argument `normalize.method = "quantile"`. However, start with real counts, derived from featureCounts instead of RSEM.
[1]: https://www.rdocumentation.org/packages/limma/versions/3.28.14/topics/voo ...
written 19 days ago by
Benn ♦ 7.8k
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Comment:
C: Row labelling heatmap in R
... I don't think it is possible with `heatmap.2`, but with `ComplexHeatmap` I know you can give labels different colors, see [here][1] for example (4th figure).
[1]: https://bioconductor.statistik.tu-dortmund.de/packages/3.1/bioc/vignettes/ComplexHeatmap/inst/doc/ComplexHeatmap.html#toc_6 ...
written 20 days ago by
Benn ♦ 7.8k
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... What I would do in such case, is to identify which genes are in that weird line. Then see what's is going on with these genes on raw count level. If that seems okay, see what average CPM values they have in the fit, search until you find some anomalies. ...
written 20 days ago by
Benn ♦ 7.8k
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... Yeah most reviewers don't even check the GEO submission, it sad but true. Did anyone contacted the authors of the paper about this? On [GEO][1] page there is an email address. If he/she won't answer it might be worth confronting the editor of the journal about this (or start a discussion on twitter) ...
written 20 days ago by
Benn ♦ 7.8k
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122
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... Most of the plots can be done in `R` (but also in other programs).
For the first you can use `barplot` from `R`.
For the second (bubble charts) you can use `ggplot2`.
The last figure looks like a network, which can be made with `RGraphviz` (or outside `R` with cytoscape). And a heatmap can be mad ...
written 21 days ago by
Benn ♦ 7.8k
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... You can accept the answer if it did what you needed. ...
written 21 days ago by
Benn ♦ 7.8k
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165
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... You need to import the matrix into R and make an `ExpressionSet` of it (https://www.rdocumentation.org/packages/Biobase/versions/2.32.0/topics/ExpressionSet). You can keep it as minimal as you want, or add stuff like pheno data and annotation. With the `ExpressionSet` you can continue with limma. ...
written 21 days ago by
Benn ♦ 7.8k
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