FAQ

How to compare the expression differences as fold changes?

You need to calculate fold change from the normalized expression levels in the diffexp.xls. You need to pay attention to interpret 'zero' expression in (all) RNA-seq data when you compare with qPCR. Zero expression means less expression than the sensitivity; it means that actual expression level might of 'zero expression by RNA-seq' might be 0.0001, 0.00001 or 0.000001, etc, when the sensitivity limit is 0.001. Therefore, the fold difference would be inaccurate when level of either case or control is zero.

What is 'spike-in based' normalized expression level?

It is relative amount of poly(A)+ transcripts to the spike-in RNAs, which were added same concentration into all samples. Therefore, it enables to detect accumulation/reduction of poly(A) transcripts per cell [Islam et al. 2011, Katayama et al. 2013], or per total RNA [Lovén et al. 2012]. In case of validation (by qRT-PCR, for example), addition and measurement of spike-in RNAs as reference is recommended.