Question: Ngs Data Normalization
1
gravatar for jack
5.4 years ago by
jack420
jack420 wrote:

Hi all,

I have RNA seq miRNA data set(NGS). it's it .txt format. I want to check that, wthether this data are normolazed or not. could sombody let me know how can I check it in R?

1    3    0    2    0    1    0
0    0    0    0    0    0    0
0    0    1    0    0    0    0
0    4    0    0    0    0    0
579    1494    401    321    65    190    39
0    0    0    0    0    0    0
0    0    0    0    0    0    0
0    0    0    0    0    0    0
0    0    0    1    0    0    0
0    0    0    0    0    0    0
0    1    0    0    0    0    0
1    1    1    1    0    0    1
41    163    42    113    25    57    14
1    1    1    0    0    0    0
44    14    25    10    6    12    5
0    0    0    0    0    0    0
0    0    0    0    0    0    0
0    0    0    0    0    0    0
0    1    0    0    0    0    0
1    0    1    0    0    0    0
0    0    0    0    0    0    0
12    10    3    2    2    5    0
ADD COMMENTlink modified 5.0 years ago by Biostar ♦♦ 20 • written 5.4 years ago by jack420
1

Could you show a few lines of the text file so we know what format the data is in? Your question is impossible to answer otherwise.

Update: Thanks, that's exactly what's needed!

ADD REPLYlink modified 5.4 years ago • written 5.4 years ago by Devon Ryan89k

Looks like data table parsed from the mirdeep2 exp files; Use EdgeR or DESEQ to normalize and do DE analysis.

check this article on NGS RNASeq normalization: http://bib.oxfordjournals.org/content/14/6/671.full

ADD REPLYlink modified 5.4 years ago • written 5.4 years ago by Rm7.8k

You can try many scaling/normalization algorithms with Normalizer: http://db.systemsbiology.net/gestalt/normalizer/

ADD REPLYlink written 5.0 years ago by JC7.7k
1
gravatar for Devon Ryan
5.4 years ago by
Devon Ryan89k
Freiburg, Germany
Devon Ryan89k wrote:

Those appear to be raw counts per miRNA. You'll want to use DESeq/edgeR/limma (see the voom() function) or one of the other standard tools to continue with the analysis.

ADD COMMENTlink written 5.4 years ago by Devon Ryan89k

I want to do some clustering with my data, but before it, i want to do quantile normalization. voom() seems doesn't support it.

ADD REPLYlink written 5.4 years ago by jack420

There's no reason for voom() to do that, it just transforms things for use in limma. If you want to do quantile normalization, just do it after importing counts with voom() (since doing quantile normalization on raw counts wouldn't make much sense).

ADD REPLYlink written 5.4 years ago by Devon Ryan89k

I don't want to use limama, I have my own algorithm. but I don't know much about NGS data, I have the above raw data and I want to normalize it and give it to my algorithm. for micoarray data, I already used quantile normalized data, but for NGS, I'm not sure what should I use . do you have any recommendation ?

ADD REPLYlink written 5.4 years ago by jack420
1

Then just perform quantile normalization on the transformed counts.

ADD REPLYlink written 5.4 years ago by Devon Ryan89k

which kind of transformation i should do before quantile normalization? is the quantile normalization is the best normalization method for NGS data ? my algorithms basically will do first clustering then dimentionality reduction and finally visulazation. I don't want to miss biological patterns due to choosing wrong normalization pattern .

ADD REPLYlink written 5.4 years ago by jack420
2

Your algorithm sounds like combat. Regarding transformation, either voom or vst (from DESeq) would work. I find the concept of a single "best" normalization method for all use cases rather simplistic.

ADD REPLYlink written 5.4 years ago by Devon Ryan89k

thanks, so what is exactly 'single "best" normalization method''?

ADD REPLYlink written 5.4 years ago by jack420
2

There isn't one

ADD REPLYlink written 5.4 years ago by Devon Ryan89k
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