Tips on integration

1. QIIME 2 Integration with EUKARYOME

1.1. Importing EUKARYOME into QIIME 2

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]'

1.2. Taxonomic analysis

With the database imported, you can now perform taxonomic analysis. The taxonomic classification can be performed with different plugins; see the basic scripts below:

Feature Classifier Plugin:


qiime feature-classifier fit-classifier-naive-bayes --i-reference-reads eukaryome_seq.qza --i-reference-taxonomy eukaryome_tax.qza --o-classifier classifier.qza

Then, you can assign taxonomy to your sequences using the trained classifier:


qiime feature-classifier classify-sklearn --i-classifier classifier.qza --i-reads your_sequences.qza --o-classification taxonomy.qza

Blast+ plugin:


qiime feature-classifier classify-consensus-blast --i-query your_sequences.qza --i-reference-reads eukaryome_seq.qza --i-reference-taxonomy eukaryome_tax.qza --p-perc-identity 0.97 --o-classification taxonomy.qza

Please note that these are basic scripts and may need to be adjusted based on your specific needs and the parameters of your data.

2. mothur Integration with EUKARYOME

Here are the following basic scripts to use with different methods in mothur:


# Wang method (default)
mothur "#classify.seqs(fasta=your_sequences.fasta, reference=mothur_SSU_v1.8.fasta, taxonomy=mothur_SSU_v1.8.tax, method=wang, cutoff=80)"

# k-nearest neighbor consensus
mothur "#classify.seqs(fasta=your_sequences.fasta, reference=mothur_SSU_v1.8.fasta, taxonomy=mothur_SSU_v1.8.tax, method=knn, cutoff=80)"

# Zap
mothur "#classify.seqs(fasta=your_sequences.fasta, reference=mothur_SSU_v1.8.fasta, taxonomy=mothur_SSU_v1.8.tax, method=zap, cutoff=80)"

Please note that these are basic scripts and may need to be adjusted based on your specific needs and the parameters of your data.

3. SINTAX Integration with EUKARYOME

To perform taxonomic analysis using SINTAX and the EUKARYOME database, use the following commands:

#converting to udb format:
vsearch --makeudb_usearch SINTAX_SSU_v1.8.fasta --output SINTAX_SSU.udb 

#assign taxonomy:
vsearch --sintax your_sequences.fasta -db SINTAX_SSU.udb -tabbedout sintax_results.txt -sintax_cutoff 0.8

Please note that these are basic scripts and may need to be adjusted based on your specific needs and the parameters of your data.

4. BLAST

First, you must format your fasta file into a BLAST database using the makeblastdb command. This command will create a nucleotide BLAST database from your fasta file:


makeblastdb -in General_SSU_v1.8.fasta -dbtype nucl -out EUK_SSU.DB

Then you can use blastn with your input sequences and the BLAST database:


blastn -query your_sequences.fasta -db EUK_SSU.DB -out results.out

Please refer to the BLAST+ manual or other relevant resources for more advanced usage.

5. DADA2

To perform taxonomic analysis using the DADA2 classifier and the EUKARYOME database, use the following commands:

library(dada2)
asv <- "input.fasta"
database <- "DADA2_EUK_ITS_v1.9.fasta"
taxa_output_file <- "taxonomy_result.csv"
taxonomy <- assignTaxonomy(asv,database, multithread =8)
write.csv(taxonomy, file = taxa_output_file)

Please refer to the DADA2 manual or other relevant resources for more advanced usage.

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