Issue with check_files.py during installation

[robertopreste]

Received via email:

Hello
I am new to Mtoolox and trying to install it. But I need some help . /MToolBox.sh

Check python version... (2.7 required)
OK.

Checking files to be used in MToolBox execution...
./MToolBox.sh: line 178: check_files.py: command not found

This is the error I find, can you please direct?

[robertopreste]

Solved as the user did not export the PATH before running MToolBox.
Follow-up questions:

I have still a few more questions to understand and try to start using the tool. If you could give me some guidance on them that would be great

1. I see that WGS reads can be used for this tool, my question is - would rna sequencing data be successfully used for this ?
2. Do I need to extract from the RNA seq file,a subset of sequencing reads that align to the mitochondrial genome or will MtoolBox do that by itself? i.e. I just provide the list of fastq files (complete raw)
3. MtoolBox uses hg19. If I have RNA seq data aligned to hg38 would that affect running the MToolBox? Can I use hg38 instad of 19 in this case?

[smukh18]

I hope I get some answers on these to know how to proceed from the developers. Please do let me know

[clody23]

Hi,

apologies for the late reply.

1. the current version of MToolBox is not meant to be used for RNA-seq. However, given the lack of splicing in mtDNA genes, the gsnap DNA-mapping mode used by MToolBox could still be used to extract the RNA variant calling. Although this is not ideal, we are aware of users doing so with their RNA-seq experiments. We are currently working on a new version of MToolBox, based on snakemake, which we plan to release in 2020, that will be flexible enough to support RNA-seq studies also.

2. No. You can just provide a BAM file with your aligned reads and MToolBox will take care of aligning mt reads on the reference sequences. Please make sure you specify the input type in the MToolBox configuration file (input_type=bam). The current MToolBox installation is based on hg19 nuclear assembly only, thus limiting the use of another genome build. However, you could create your own GSNAP indexes and make sure to overwrite the values of the MToolBox variables defined in the setup file (https://github.com/mitoNGS/MToolBox/blob/master/MToolBox/setup_example.sh) or in the MToolBox configuration file (https://github.com/mitoNGS/MToolBox/blob/master/test_rCRS_config.sh )
   e.g.,

gsnapdb=/your/dir/with/gsnap_db/
mtdb_fasta=chrM.fa
hg19_fasta=hg38RCRS.fa
mtdb=chrM
humandb=hg39RCRS.fa

3. No, if you want to keep hg19 assembly as reference sequence, MToolBox will take care of extracting the files from the BAM input file and realign. If you want to use hg38, please see point 2.

Best regards,

Claudia