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qiita_pet/support_files/doc/source/processingdata/qp-fastp-minimap2.rst
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Adapter and host filtering | ||
========================== | ||
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At the end of August 2023, we discovered that the parameters used by | ||
qp-fastp-minimap2 did not trigger application of adapter filtering. By default, | ||
fastp performs autodetection of adapters and filtering for single-end data. By | ||
default, fastp does not perform these operations on paired-end data. This behavior | ||
was not expected by us. It was discovered when manually assessing replicated | ||
sequences, which on examination by BLAST against NT reported to be adapters. | ||
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Adapter filtering for paired-end data with fastp requires specifying either the | ||
exact adapters to remove (i.e., no autodetection), or to explicitly specify “--detect_adapter_for_pe”. Qiita previously indicated to users that the | ||
qp-fastp-minimap2 plugin was performing adapter autodetection and filtering. | ||
However, because this flag was not specified, that behavior did not occur. | ||
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In the metagenomic dataset the adapters were discovered in, we observed a few | ||
sequences with high replication, which assignments to a few genomes in RS210. | ||
The coverage of those genomes, using all metagenomic short reads, was constrained | ||
to very specific regions. The replicated sequences exhibited high identity to | ||
known adapters. As such, we suspect the replicated sequences we observed were | ||
adapters. We suspect the observed genomes either suffer from adapter contamination | ||
themselves, or the constructs used in the samples we examined were derived from | ||
real organisms. Although we cannot differentiate this definitively in the data | ||
we examined, in either case these short reads are likely artifactual. | ||
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For the dataset we examined, removal of these false positives was important | ||
for the biological interpretation of the results. However, whether the removal | ||
is important likely depends on the dataset and question. | ||
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qp-fastp-minimap2 has been updated to perform adapter filtering on paired-end data. | ||
The fastp autodetection is compile-time limited to `the first 256k sequences <https://github.com/OpenGene/fastp/blob/7784d047fdf0a8df4211967156f5c97920c6d2e8/src/evaluator.cpp#L410-L417>`_. | ||
Because of this, we opted for a more conservative approach of not relying on | ||
autodetection and instead we now test all adapters that fastp is aware of. Specifically, | ||
we now provide fastp a known adapters FASTA which is a serialized representation | ||
of their `known adapter list <https://github.com/OpenGene/fastp/blob/7784d047fdf0a8df4211967156f5c97920c6d2e8/src/knownadapters.h#L11>`_. | ||
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The new command is named: `Adapter and host filtering v2023.12`. |
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