Question: rRNA remove from RNAseq data
0
gravatar for Peter
15 months ago by
Peter0
Peter0 wrote:

Hi everyone,

I mapped human RNAseq raw data to Refseq rRNA sequences using bwa, it turned out that 20% raw reads mapped to rRNA sequences. Is it normal? But I have used a kit to remove rRNA including cytoplasmic(5s, 5.8s, 18s, 28s) and mitochondrial(12s, 16s) ribosomal RNA. I found that the most mapped reads belong to 45S pre-ribosomal RNA sequences. I have no idea about this. Someone could help me! Any suggestion is welcome!

Thanks!

rna-seq • 1.7k views
ADD COMMENTlink modified 15 months ago by ATpoint24k • written 15 months ago by Peter0
1

You could use riboPicker to identify and remove rRNA reads from the fastq files. I am not sure if SILVA databases include the pre-rRNA sequences though.

ADD REPLYlink modified 15 months ago • written 15 months ago by Sej Modha4.5k
1
gravatar for firatuyulur
15 months ago by
firatuyulur290
firatuyulur290 wrote:

I have used SortMeRNA before and worked very well for me.

ADD COMMENTlink written 15 months ago by firatuyulur290
1
gravatar for ATpoint
15 months ago by
ATpoint24k
Germany
ATpoint24k wrote:

Do you plan to assemble a transcriptome? If not, you don't have to remove them. When assigning reads to feature of a GTF, they will not be counted anyways. 20% does not sound too bad. If you did not use a kit at all, it would be like 95%, so as long as you get sufficient reads mapping on target for your analysis, I would not care about them. Still as a remark, BWA is not a splice-aware aligner. You might consider using HISAT2 or STAR for RNA-seq alignments.

ADD COMMENTlink modified 15 months ago • written 15 months ago by ATpoint24k

This sounds like a good suggestion, it is not clear if OP is carrying out standard RNASeq analysis or something else e.g. metatranscriptomics.

ADD REPLYlink written 15 months ago by Sej Modha4.5k

thank you! I only see the rRNA rate in my fastq file using BWA for mapping to refseq rRNA sequences. I just want to know the efficiency of the kit used. I don't know why 45S pre-rRNA is not removed. 20% accounts for about 6G of data, which I think is a big waste. I want to reduce this waste. I am carrying out standard RNASeq analysis.

ADD REPLYlink written 15 months ago by Peter0
0
gravatar for Antonio R. Franco
15 months ago by
Spain. Universidad de Córdoba
Antonio R. Franco4.1k wrote:

There is a program named BBSplit that you can use to filter and remove your undesired sequences

ADD COMMENTlink written 15 months ago by Antonio R. Franco4.1k
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