Reference-based chimera detection

1. UCHIME reference-based chimera detection

Using the EUKARYOME database for the reference-based chimera filtering process in USEARCH or VSEARCH (–uchime_ref):

 

vsearch --uchime_ref your_sequences.fasta 
--db General_EUK_ITS_v1.8.fasta 
--nonchimeras nonchimeras.fasta

2. QIIME 2 reference-based chimera detection

Initially, you should change the format to the “.qza” format and import the files into QIIME 2:

 

Importing the taxonomy file:


qiime tools import --type 'FeatureData[Taxonomy]' --input-path QIIME2_EUK_SSU_v1.8.tsv --output-path eukaryome_tax.qza

Importing the sequence file:


qiime tools import --input-path QIIME2_EUK_SSU_v1.8.fasta --output-path eukaryome_seq.qza --type 'FeatureData[Sequence]'

QIIME 2 uses the VSEARCH plugin for reference-based chimera detection. Here’s how you can use it: 

qiime vsearch uchime-ref 

--i-table table.qza 

--i-sequences your-seqs.qza 

--i-reference-sequences eukaryome_seq.qza 

--o-nonchimeras  nonchimeras.qza

It takes as input:

  • table.qza: The feature table was obtained from the sequence quality control step.

  • your-seqs.qza: The representative sequences obtained from the sequence quality control step.

  • eukaryome_seq.qza: A reference database of chimeric sequences

  • nonchimeras.qza: The output file where non-chimeric sequences will be stored

 

3. Mothur reference-based chimera detection

mothur provides the chimera.uchime command for reference-based chimera detection:

mothur "#chimera.uchime(fasta=your_sequences.fasta, reference=mothur_EUK_SSU.v1.8.fasta)"

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