Normalisation before RUVg and RUV2?
1
0
Entering edit mode
Pauly Lin ▴ 150
@pauly-lin-7537
Last seen 8.4 years ago
University of New South Wales, Australia

Hi!

I want to use the RUVSeq package to remove the unwanted variation in my RNA-seq data. Should I apply the function RUVg on raw counts or normalised counts (UQ normalised or TMM normalised)?

Similarly, I want to use the RUV2 function in the ruv package to remove the unwanted variation in my micro-array data - should I apply the function on unnormalised data or normalised data?

Cheers

Paul

ruvseq ruv • 2.6k views
ADD COMMENT
3
Entering edit mode
davide risso ▴ 950
@davide-risso-5075
Last seen 3 days ago
University of Padova

Hi Paul,

 

in the RUVSeq vignette we do scale for UQ prior to compute the RUVg factors. But on our data, RUVg without scaling leads to similar results. The only thing to keep in mind is that if you do scale before computing the RUV factors, you then need to remember to put an offset in the model via, e.g., the calcNormFactors function of edgeR. This is detailed in the vignette. If you decide not to scale, then in principle the RUV factors will account for sequencing depth too, so you may want to force edgeR's offset to 0.

As for RUV-2, I believe the results are similarly robust to prior normalization. I would apply RUV-2 on the normalized data, but I think you could get reasonable results on the raw data, as well.

Best,

davide

ADD COMMENT
0
Entering edit mode

Thanks, Davide!

Paul

 

ADD REPLY

Login before adding your answer.

Traffic: 769 users visited in the last hour
Help About
FAQ
Access RSS
API
Stats

Use of this site constitutes acceptance of our User Agreement and Privacy Policy.

Powered by the version 2.3.6