Question: Help required for Bacterial RNAseq data
0
gravatar for bioinforesearchquestions
19 months ago by
United States
bioinforesearchquestions200 wrote:

Hi friends,

I am going to 8 bacterial bulk RNAseq samples. I am interested in doing following bioinformatics analyses for the 12 samples.

a) Trimming of the reads

b) Alignment of the RNA-seq reads to the reference bacterial genome

c) ID and construction of splice-junctions

d) Reports of known transcripts with annotation and abundance

e) Report of novel transcripts and abundance

f) Testing differential expression

g) GO and Kegg annotation and enrichment analysis

h) Also, DeNovo Assembly

1) For mammalian, I used the following softwares Tophat, cufflinks, cuffmerge, cuffquant, and cuffdiff? 2) Is there any separate tools/pipelines for bacterial RNAseq data? 3) What are tools for GO, Kegg and enrichment analysis? 4) Which denovo assembly tool is recommended for bacteria?

gene expression rnaseq • 824 views
ADD COMMENTlink modified 18 months ago by Biostar ♦♦ 20 • written 19 months ago by bioinforesearchquestions200
2

For # 1 since you would not expect splicing to be an issue you could use any aligner (I recommend BBMap suite for trimming/alignment).
#2 there are dedicated pipelines available for bacterial RNAseq. Rockhopper is a web based one. Search Entrez for others.
#3 You should be able to use GeneSCF (Gene Set Clustering based on Functional annotation (GeneSCF) ).
#4 SPAdes is about the best bacterial genome assembler.

ADD REPLYlink modified 19 months ago • written 19 months ago by genomax65k

Thanks, Genomax for your recommendations. I checked SPAdes website and I found there is a separate "De Novo RNAseq Assembler" called "rnaSPAdes".

For mammalian RNAseq data, I started with FASTQ files --> BAM/SAM from Tophat --> raw read count from HTseq-count or GTF file from Cufflinks --> merged GTF from Cuffmerge --> differential outpu from Cuffdiff.

For bacterial RNAseq data, when I don't have reference genome. Using rnaSPAdes assembler, I will get the output file in FASTA format. But with that FASTA file how do I address (d, e, f, g, h)?

ADD REPLYlink modified 19 months ago • written 19 months ago by bioinforesearchquestions200
1

Take a look at Trinity, which does everything that you want: https://github.com/trinityrnaseq/trinityrnaseq/wiki

Rockhopper, mentioned by genomax is good. It runs from your local disk as a JAVA jar file, too. Afterwards, you can infer functionality of the identified RNA transcripts via BLASTx

ADD REPLYlink written 18 months ago by Kevin Blighe41k
1

Thanks, Kevin. I will first run trinity and see what are all the output files I am going to get.

ADD REPLYlink written 18 months ago by bioinforesearchquestions200
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