problem in Abundance Estimation using RSEM

Question

[bilal.sarwar]

Hello all I am a new user of Linux OS. Recently I received my paired-end RNA seq data. I used trinity for Denovo assembly successfully. the script was....

/home/bilal/Denovo/trinityrnaseq-2.2.0/Trinity --seqType fq \ --left /home/bilal/Denovo/left.fq \ --right /home/bilal/Denovo/right.fq \ --SS_lib_type RF \ --max_memory 58G \ --CPU 10 \ --output /home/bilal/Denovo/trinity_AssebleSeq

my next object is abundance estimation Using RSEM. the code I used for abundance estimation .......

/home/bilal/Denovo/trinityrnaseq-2.2.0/util/align_and_estimate_abundance.pl \ transcripts /home/bilal/Denovo/trinity_AssebleSeq/Trinity.fasta \ --seqType fq --left /home/bilal/Denovo/files/control1.fastq \ --right /home/bilal/Denovo/files/control2.fastq \ --est_method RSEM \ --aln_method bowtie \ --trinity_mode \ --prep_reference \ --output_dir /home/bilal/Denovo/RSEM

it showed following error

*bilal@Plant-Genomics:~/Denovo/trinityrnaseq-2.2.0/util$ sh RSEM2.sh --transcripts: not found --left: not found --trinity_mode: not found*

I give the file path correctly, the trinity OK. please help me how to fix this error

thanks

Answer 1

[WouterDeCoster]

I'm not sure since your code might be formatted when posting here, but shouldn't that be align_and_estimate_abundance.pl --transcripts? You didn't have the --.

[bilal.sarwar]

thanks for your response @WouterDeCoster

##fixed now above mentioned

when i run the align_and_estimate_abundance.pl with the command

/home/bilal/Denovo/trinityrnaseq-2.2.0/util/align_and_estimate_abundance.pl --seqType fq \ --transcripts /home/bilal/Denovo/test/trinity_AssebleSeq/Trinity.fasta \ --left /home/bilal/Denovo/test/reads.left.fq \ --right /home/bilal/Denovo/test/reads.right.fq \ --SS_lib_type RF \ --est_method RSEM \ --aln_method bowtie2 \ --trinity_mode \ --prep_reference \ --output_dir /home/bilal/Denovo/rsm

it throws the following error

ERROR, cannot find rsem-calculate-expression in PATH setting: /home/bilal/bin:/home/bilal/.local/bin:/usr/local/sbin:/usr/local/bin:/usr/sbin:/usr/bin:/sbin:/bin:/usr/games:/usr/local/games:/snap/bin:/home/bilal/Denovo/trinityrnaseq-2.2.0/util

Please be sure bowtie and express are installed and the utilities samtools bowtie-build bowtie rsem-calculate-expression are available via your PATH setting. ................................................ please help me

[WouterDeCoster]

The script wants to execute rsem-calculate-expression but can't find it, because it's not installed somewhere where your operating system is looking for executables/scripts.

The variable $PATH (you can check this one with typing echo $PATH in your terminal) specifies in which directories scripts will be looked for. So in this case you can a) add the directory containing rsem-calculate-expression to your $PATH or b) make sure that a copy or link to rsem-calculate-expression is present in a directory which is in $PATH

I assume the installation instructions will contain something about this. This would also have the effect that you don't have to specify the complete path to a script with every command.

Let me know if this wasn't completely clear.

[bilal.sarwar]

Thanks a lot @WouterdeCoster for your valuable suggestion. it's fixed NOW :) GOD bless you thanks