Submitting a metagenome bin

The submission of the bin can be done only using webin-cli.

Again, We will need to create a manifest file, which contains the metadata for our read data and references the assembly fasta file that should be submitted.

First, we create another directory:

mkdir -p /mnt/submission/bins/bins

Now, choose one of the bin files from the metabat results in /mnt/WGS-data/megahit_out/metabat/final.contigs.fa.metabat-bins-*/ and copy it to the metabat folder:

cp <your_bin> /mnt/WGS-data/megahit_out/metabat/

Then, we gzip the bin file, since it needs to be zipped for submission:

gzip /mnt/WGS-data/megahit_out/metabat/bin.*.fa

And change to our submission directory:

cd /mnt/submission/bins/bins

Create the manifest file

We will use this short manifest template for our submission (almost the same as the assembly except for the ‘binned metagenome’ and different fasta file ofr course), fill in YOUR values for STUDY, SAMPLE and RUN accession, be careful to fill in your BINNED SAMPLE ACCESSION, not the environmental sample accession (webin-cli won’t complain if you take the wrong one):

STUDY   TODO
SAMPLE   TODO
RUN_REF   TODO
ASSEMBLYNAME   TODO
ASSEMBLY_TYPE   binned metagenome
COVERAGE   20
PROGRAM   MEGAHIT
PLATFORM   ILLUMINA
MOLECULETYPE   genomic DNA
FASTA  /mnt/WGS-data/megahit_out/metabat/bin.<YOUR_BIN>.fa.gz

Create a file named manifest and fill it with the content above - fill the fields marked with TODO with the appropriate content. Then continue with the next step.

Validating the bin submission

Before actually submitting, we are validating our manifest file. To do so, we use the option -validate in our call of webin-cli. Also, make sure, to use the -test flag to submit to the ENA test server. We also use -context=genome since we are submitting an assembly. Other options are your -username, -password and the path to the -manifest file:

cd /mnt/submission/bins/bins
java -jar ~/webin-cli-6.10.0.jar -username=$ENA_USER -password=$ENA_PWD -context=genome -manifest=manifest -validate -test

When everything was successfully validated, you should get a message like:

INFO : The submission has been validated successfully.

Submit the bin

Now, that our bin submission is validated successfully, we can go on with the submission. Just replace the -validate flag by -submit in the webin-cli call. Do NOT remove the -test flag:

cd /mnt/submission/bins/bins
java -jar ~/webin-cli-6.10.0.jar -username=$ENA_USER -password=$ENA_PWD -context=genome -manifest=manifest -submit -test

If everything works fine, you should receive a message like:

INFO : The TEST submission has been completed successfully. This was a TEST submission and no data was submitted. The following analysis accession was assigned to the submission: ERZ14243535

Great! Now let’s go on by adding a MAG to this submission.

References

ENA - Submitting Binned Metagenome Assemblies https://ena-docs.readthedocs.io/en/latest/submit/assembly/metagenome/binned.html