Question: Prokaryotic Differential expression analysis- RNA seq data and software
0
gravatar for zh.khodadadi
2.5 years ago by
zh.khodadadi20
zh.khodadadi20 wrote:

Hi every body I want to analyze(Differential expression ) my rna seq data of bacteria single end read,50 bp, Illumina HiSeq 2000. Can I use tuxedo software? What is the pipeline of Differential expression analysis for bacteria rna seq data? thanks a lot

ADD COMMENTlink modified 2.5 years ago • written 2.5 years ago by zh.khodadadi20
2

Since you don't need to account for splicing you could use any NGS aligner and count the reads that are aligning to the genes using an annotation file (GTF/GFF). This can then be followed by one of the popular DE analysis programs DESeq2/edgeR etc.

An online pipeline is mentioned in this thread (if that is of interest): Bacterial RNA-seq analysis

ADD REPLYlink modified 2.5 years ago • written 2.5 years ago by genomax65k

Dear genomax2, Hi

Is this pipeline helpful for bacteria?

FastQC --> Trimmomatic/Cutadapt --> STAR --> Samtools --> featureCounts/HTseq-count --> DESeq2

ADD REPLYlink modified 2.5 years ago • written 2.5 years ago by Farbod3.2k

STAR can output sorted bam, so I'm not sure why you add samtools in there ;)

ADD REPLYlink written 2.5 years ago by WouterDeCoster38k

Hi,

I have used it from other Biostars posts, so it should be :

FastQC --> Trimmomatic/Cutadapt --> STAR --> featureCounts/HTseq-count --> DESeq2

ADD REPLYlink written 2.5 years ago by Farbod3.2k
4

I am partial to BBMap suite so I would do

FastQC --> bbduk (from BBMap) --> bbmap (from BBMap)  --> featureCounts --> DESeq2

BBMap will output bam files directly (as long as samtools is available in $PATH, but you would need to sort them). featureCounts will sort BAM's automatically, in case you do not want to use samtools.

ADD REPLYlink written 2.5 years ago by genomax65k

Seems valid, but as @genomax2 already wrote a spliced aligner isn't really necessary (because no splicing in bacteria), and I also would run fastQC after trimming to see how successful it was.

ADD REPLYlink written 2.5 years ago by WouterDeCoster38k

Dear zh.khodadadi, Hi

Do you have any reference genome for your bacteria of interest?

~ Best

ADD REPLYlink written 2.5 years ago by Farbod3.2k

thanks all I am new in NGS why I dont use Tuxedo pipeline?

ADD REPLYlink written 2.5 years ago by zh.khodadadi20

My bacteria has reference genome and I study some posts like that genomax2 says.

ADD REPLYlink written 2.5 years ago by zh.khodadadi20

Please use ADD REPLY to answer to earlier comments, as such this thread remains logically structured and easy to follow.

ADD REPLYlink written 2.5 years ago by WouterDeCoster38k

ok thanks for this guidance

ADD REPLYlink written 2.5 years ago by zh.khodadadi20
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