Inputs of MetaPUF

This workflow offers the users to integrate publicly available metagenomics, metatranscriptomics and metaproteomics datasets on PRIDE and MGnify portals.

The workflow requires the following inputs in the config.proteomics.yaml file:

Mandatory inputs

  • Study : Under the parameters section, which takes in European Nucleotide Archive (ENA) secondary study accession: starts with (ERP|DRP|SRP) followed by six digits, e.g. ERP124921.

  • Input_dir/Pride_id: Under the parameters section, one (only one) of them needs to be enabled. If you have the MS raw files available in your local machine, you can enable the Input_dir to give the path of the raw files as an input directory, or provide the PRIDE accession number if the raw files are publicaly available in PRIDE archive.

We suggest downloading the raw files to your local machine and do the further analysis. Since the raw files downloading process usually takes a lot of time, and it will need to download from the beginning if the pipeline breaks when running.

  • Metadata: Under the raws section, it is .csv file (defalut name: sample_info.csv) which should be a relationship table between the metaproteomics raw files and the metagenomics and/or metatranscriptomics assemblies, this is mandatory as well because the data identifiers between PRIDE, MGnify and ENA are not related.

Please make sure that the input metagenomics and/or metatranscriptomics assemblies and metaproteomics data are from the same samples.

  • .csv file: which is the file mentioned in the Metadata parameter contains all the sample information, please put this .csv file and the config.proteomics.yaml file in the same folder: config.

An example for this file:

Sample

Raw file

Raw file URLs

Sample Accession

Assembly

S6

S6.raw

https: //ftp.pride.e bi.ac.uk/prid e/data/archiv e/2017/07/PXD 005780/S6.raw

ERS1509315

ER Z1669330

You should leave the columns of the Raw file URLs as blank if you have the raw files locally, however the header is still needed.

Other input parameters

The input parameters for the pipeline are all set in the config.proteomics.yaml file and you can change them based on your own preferences. Some instructions regarding the parameters:

  • Version : The version of MGnify analysis. This accepts either “4.1” or “5.0” as string values. The default is “5.0”.

  • Db_size : Size of the proteins sequence database in bytes. This is mentioned in our manuscript, when the reference protein database is too large, the pipeline will apply a tree traversal algorithm to dynamically generate multiple Reference Search Databases based on the clusters of the samples. The default is 1073741824 (integer data type).

  • outputdir : Directory path to save the output

There are some other output folders for searchgui and peptideshaker, the users can change the name themselves. Also, the parameters in the searchgui section needs to be changed based on the Mass spectrometry analysis.