Question: Differential Gene Expression Analysis By Rna-Seq
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gravatar for nayaganviji
6.5 years ago by
nayaganviji20
nayaganviji20 wrote:

I am planing to find out differential gene expression between two samples (infected and non infected RNA-seq data of avian species from 454 GSFLX+ PLATFORM), but there is no reference genome available. So I am planning to perform a De No transcriptome assembly with Newbler. How can I determine the number reads assembled to particular gene (I have only contigs from the assembly) for statistical significance testing. Are any normalization procedures required? Please suggest some common software for this analysis.

Thanks

rna-seq • 3.0k views
ADD COMMENTlink modified 3.8 years ago by Biostar ♦♦ 20 • written 6.5 years ago by nayaganviji20
2

Have you searched the forum? A similar question has been asked: http://www.biostars.org/post/show/17611/ Also, are you familiar with R and bioconductor? There are some powerful tools between those two that can answer the questions you are asking.

ADD REPLYlink written 6.5 years ago by seidel6.8k
3
gravatar for JC
6.5 years ago by
JC7.7k
Mexico
JC7.7k wrote:

When you don't have a reference genome, makes sense to assembly your sequences to get the transcripts, one strategy I have seen before is to assemble all your libraries in one run to get the better representation of the transcriptome and then map back your reads per library to get the counts per condition. Then you can use R:Bioconductor edgeR or DESeq to deal with normalization and differentially expressed transcripts.

ADD COMMENTlink modified 6.5 years ago • written 6.5 years ago by JC7.7k

This is what Brian Haas recommends when he teaches. The Trinity folks put together some documentation on how to do it with Trinity here. You don't have to use Trinity and EdgeR but the logic is all laid out.

http://trinityrnaseq.sourceforge.net/analysis/diff_expression_analysis.html

ADD REPLYlink written 5.3 years ago by Michele Busby1.9k
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