Dear edgeR users and developers，

I have few questions about edgeR when recently I use it for 454  
pyrosequencing data:

1. prior.n
     According to users' manual, we may not use too low prior.n in  
moderated tagwise dispersion approach. But in my dataset, there are  
more than 15 samples in each comparison group and the freedom is  
larger than 30. prior.n <- estimateSmoothing(d) gives 0.0005329. So I  
am wondering if I could use 0.0005329 since I have rather big number  
of samples in each group. Or I should adjust prior.n into 10 according  
to the manual's suggestion.

2. TMM
     I am not sure if this is also applicable to 454 microbiota data.  
I suppose I should do TMM normalization as well since the  
normalization factors from my samples have a big variation (f is from  
0.41 to 4.58). Is that right?

3. p_value
     According to your experience, is it reasonable and reliable to  
use p_value < 0.05 as significance criteria? or only <0.01 can be  
reliable.

I am a new users in this package and hope you may give some  
suggestions. Many thanks!

Ying Ye





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