Dear Michael,
thanks for your instantaneous reply!
> I hope I can bother you with some questions about your package:
> 1. when analysing for let's say two variables (e.g. ~ type +
> condition), will this be a difference to change the order of the
> variables (~ condition + type)? Probably this will do a blocking
> and hence order is important here similar to edgeR?
>
>
> it makes no difference except that the results() function with no
> arguments will automatically extract results for the last variable in
> the design formula, and for the last level of this variable over the
> first level if this variable is a factor. But otherwise, no it
> doesn't make a difference if you are using the arguments of results()
> to specify which results tables to construct.
>
> 2. For more than 2 variables (e.g. ~type + condition + time) would
> this be meaning blocking will also be done for the further
> variables (e.g. type for time)? If I intend no blocking I probably
> need to do ~ 0 + type + condition + time?
>
>
> Yes, adding variables "accounts for" or "controls for" other effects,
> whether there are 2 variables, 3, 4 etc.. We do not recommend using
> the "0" in design formula for DESeq2, because by default we shrink
> non-intercept effects (see the description of betaPrior in ?DESeq),
> but we want to have an unshrunken intercept. Can you say more what you
> mean by "intend no blocking", in my mind, blocking is a property of
> the experiment.
>
My understanding from working with the edgeR package is, that you can
consider paired samples or batch effects when comparing two or more
treatments. Accounting for paired samples seems to be a specific case of
"blocking" in experimental design. In edgeR doing ~type+condition I can
retrieve the 'condition' comparison including the correction for the
paired effect 'type'. Is this also true for DESeq2? And if so, does the
order matter in the formula e.g.: ~type+condition the same as
~condition+type?
>
> 3. Is it also possible to study nested effects and other
> interactions (e.g. condition:time)?
>
>
> Yes, you can include interaction terms in the design formula. If you
> are using DESeq2 v1.2 you can pull out the effects by 'name' argument
> to results(), using a name in resultsNames(dds). In DESeq2 >= 1.3, I
> have added instructions and examples to the ?results manual page, but
> still need to add a section to the vignette on interaction terms.
>
Cool!
Thanks a lot,
Claudia
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