Question: RPKM with bowtie output
0
gravatar for flogin
13 months ago by
flogin250
Brazil
flogin250 wrote:

Hello guys,

There is a way to evaluate RPKM direct with bowtie 2 output (e.g. bam or sorted bam)? like an R package or some scripts to do this?

count bowtie rna rpkm • 490 views
ADD COMMENTlink modified 12 months ago by anoo0 • written 13 months ago by flogin250
2

Is this RNA-seq? Are you aware bowtie2 is not splice-aware? What organism? Do you have a GTF file or list of reference peaks (if not RNA-seq)?

ADD REPLYlink modified 13 months ago • written 13 months ago by ATpoint41k

It's a mapping of RNA-seq against a set of contigs, I just have the contigs, rna-seq files and bam files.

ADD REPLYlink written 13 months ago by flogin250
4
gravatar for swbarnes2
13 months ago by
swbarnes29.2k
United States
swbarnes29.2k wrote:

RSEM will take a bam file as input and return raw counts, RPKM and TPM

ADD COMMENTlink written 13 months ago by swbarnes29.2k

thanks swbarnes2, I'll check this.

ADD REPLYlink written 13 months ago by flogin250

Hello, Can you please provide the steps used for calculating expression using RSEM, after building the reference. The script is throwing errors.

ADD REPLYlink written 12 months ago by anoo0

Please read the manual and then come back with more specific questions while opening a new question, providing the necessary details.

ADD REPLYlink written 12 months ago by ATpoint41k

The script is throwing errors.

How do you expect us to help if you don't tell us what's going wrong?

ADD REPLYlink written 12 months ago by WouterDeCoster44k

I used the same commands given in the manual, with the BAM files I have,

software/RSEM-1.2.25/rsem-calculate-expression -p 8 --paired-end \ --bam \ --estimate-rspd \ --append-names \ --output-genome-bam \ exp/LPS_6h.bam \ ref/mouse_ref exp/LPS_6h

Error:

Warning: The SAM/BAM file declares less reference sequences (955) than RSEM knows (41078)! Please make sure that you aligned your reads against transcript sequences instead of genome. RSEM can not recognize reference sequence name 1!

"rsem-parse-alignments /home/user/data/RSEM/Rat /home/user/data/RSEM/Rat.temp/Rat /home/user/data/RSEM/Rat.stat/Rat /home/ec2-user/data/Rat/*.bam 3 -tag XM" failed! Plase check if you provide correct parameters/options for the pipeline!

ADD REPLYlink written 12 months ago by anoo0
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