JGI Metagenome Assembly in KBase

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A KBase module generated by the KBase SDK.

This module implements the JGI metagenome assembly pipeline in KBase. This implementation is still pending some testing and refinement, but works as follows:

Current limitations

This version only works on Paired End Illumina reads. single end reads or reads from other platforms will likely fail in strange or spectacular ways.

Input:

A single paired end reads library composed of sequences read from an Illumina machine.

Output:

An assembled set of contigs, as an Assembly object, along with a brief report on the results.

Computational Steps:

1. BBTools RQCFilter (see the KBase-wrapped BBTools or the App Catalog page for details). This produces a library of reads that pass an automated quality control, removes reads that match a standard set of contaminants, trims adapters and linkers.
2. BFC version r181 (paper, Github) This corrects sequencing errors on short reads generated by Illumina sequencing technology. Uses parameters -1 -k 21 -t 10
3. seqtk (Github) Seqtk is a toolkit for managing FASTA and FASTQ files. It's used here to remove orphan single end reads from the paired end library, after filtering and error correction. Used with the dropse subcommand.
4. SPAdes version 3.12.0 (paper, website) This does the work of assembling the reads. Uses parameters --only-assembler -k 33,55,77,99,127 --meta -t 32 -m 2000. Therefore, this uses metaSPAdes with k-mers values of 33, 55, 77, 99, and 127. A caveat here is that if your reads (after filtering and correcting) have an average length that's less than any of those k-mer values, that k-mer value won't be used. E.g., if your corrected reads have an average length of 100, the k=127 pass will be skipped.
5. BBTools stats version 38.22 (website) Two more tools are used from BBTools at the end. The first is stats.sh to build the assembly statistics. Run with flag format=6.
6. BBTools bbmap version 38.22 BBMap is finally used at the end to build a coverage map of reads mapped onto the finalized assembly, and generate coverage statistics. Uses default parameters, except for the flag ambiguous=random.

Next steps

1. Add flexibility to auto-detect and adapt to single end reads. This mainly means doing the detection at app start, and adjusting a few pipeline parameters in the process.
2. Add an option to use Quast to show a deeper report on assembly quality.
3. Add an option to use Qualimap to show a deeper report on the generated reads mapping to assembly.

Technical notes

• Updating versions - update the versions of SPAdes or BBTools by modifying the values in pipeline.cfg. Those should match the strings in the name of the downloaded objects, so you'll need to be careful with that. Note this only modifies the version of the tool being run and the text in reports - any documentation text that refers to the version (like above) needs to be adjusted by hand.
• RQCFilter failures - RQCFilter runs as a local method (see the KBase SDK docs). Running tests of this pipeline that call out to RQCFilter in a way that has all of its required reference data is tricky, so expect a total of one test failure if you run this module's test suite locally.