BIP Platform wrapper scripts for standardized execution of nf-core/ampliseq
This repository provides helper scripts and usage instructions to simplify processing of raw amplicon data into cleaned, standardized ASV tables.
It ensures consistent sample naming, reproducible parameterization, and automated cleanup of intermediate files.

These scripts are also available at:
/work_ikmb/ikmb_repository/shared/microbiome/RUN_AMPLISEQ/

cd /work_ikmb/sukmb276/Microbiome/clean_data_from_dada2/Runs_v.1.10_Fungi
mkdir XXMonthXXX
cd XXMonthXXX

Use the helper script to create a standardized samplesheet.tsv based on raw FASTQ filenames:

/work_ikmb/ikmb_repository/shared/microbiome/RUN_AMPLISEQ/generate_samplesheet.sh \
  --input_dir /path/to/raw_data

We recommend running the pipeline inside a tmux session to avoid job interruptions:

tmux new-session

# Example 1: ITS2 fungi (paired-end)
/work_ikmb/ikmb_repository/shared/microbiome/RUN_AMPLISEQ/run_ampliseq.sh \
  --input samplesheet.tsv \
  --outdir results \
  --primers its2 \
  --db unite

# Example 2: 18S eukaryotes (paired-end)
/work_ikmb/ikmb_repository/shared/microbiome/RUN_AMPLISEQ/run_ampliseq.sh \
  --input samplesheet.tsv \
  --outdir results \
  --primers 18s \
  --db pr2

# Example 3: 18S eukaryotes (single-end, forward only)
/work_ikmb/ikmb_repository/shared/microbiome/RUN_AMPLISEQ/run_ampliseq.sh \
  --input samplesheet.tsv \
  --outdir results \
  --primers 18s \
  --db pr2 \
  --single_end

For GMBC V3-V4 processing, use the dedicated script in this repository:

bash run_ampliseq_gmbc_v3v4.sh

This script uses:

• nf-core/ampliseq 2.16.1
• V3-V4 primers (CCTACGGGAGGCAGCAG / GGACTACHVGGGTWTCTAAT)
• --db gtdb
• --outdir results_filtered
• Filtering settings (--min_samples 2, --min_frequency 10)

Why this is needed for GMBC:

• The generic wrapper may skip downstream QIIME2 steps in some modes.
• The GMBC script keeps the QIIME2 branch active (no --skip_qiime* flags), which is required to produce phylogeny outputs.
• This is the expected path when you need phylogenetic tree generation for downstream analyses.

Input files expected in the working directory:

• samplesheet.tsv
• metadata.tsv
• custom.config (already provided in this repository and points to ampliseq_custom.config)

For GMBC 18S processing, use the dedicated script in this repository:

bash run_ampliseq_gmbc_18s.sh

This script uses:

• nf-core/ampliseq 2.16.1
• 18S primers (TTAAARVGYTCGTAGTYG / CCGTCAATTHCTTYAART)
• --db pr2
• --outdir results_filtered_18s
• Filtering settings (--min_samples 2, --min_frequency 10)

Why this is needed for GMBC:

• It mirrors the GMBC V3-V4 dedicated execution model for consistency.
• It keeps the QIIME2 branch active (no --skip_qiime* flags), which is required for phylogeny outputs.
• It applies 18S-specific taxonomy levels and uses the 18S custom config.

Input files expected in the working directory:

• samplesheet.tsv
• metadata.tsv

After the pipeline finishes, clean the output and extract seqtab with:

cd /work_ikmb/sukmb276/Microbiome/clean_data_from_dada2/Runs_v.1.10_Fungi/XXMonthXXX

/work_ikmb/ikmb_repository/shared/microbiome/RUN_AMPLISEQ/prepare_seqtab.sh

For 18S rRNA analysis with single-end reads:

/work_ikmb/ikmb_repository/shared/microbiome/RUN_AMPLISEQ/run_ampliseq.sh \
  --input samplesheet.tsv \
  --outdir results \
  --primers 18s \
  --db pr2 \
  --single_end

Note: For 18S analysis:

• Paired-end mode: Uses ampliseq_custom_18s.config (with tryRC=TRUE for proper taxonomic assignment)
• Single-end mode (default): Uses ampliseq_custom_18s_no_tryRC.config (without tryRC=TRUE)

/work_ikmb/ikmb_repository/shared/microbiome/RUN_AMPLISEQ/run_ampliseq.sh \
  --input samplesheet.tsv \
  --outdir results \
  --primers its2 \
  --db unite \
  --resume

/work_ikmb/ikmb_repository/shared/microbiome/RUN_AMPLISEQ/run_ampliseq.sh \
  --input samplesheet.tsv \
  --outdir results \
  --primers its2 \
  --db unite \
  --multiple_sequencing_runs

To explore script options:

/work_ikmb/ikmb_repository/shared/microbiome/RUN_AMPLISEQ/run_ampliseq.sh --help
/work_ikmb/ikmb_repository/shared/microbiome/RUN_AMPLISEQ/generate_samplesheet.sh --help
/work_ikmb/ikmb_repository/shared/microbiome/RUN_AMPLISEQ/prepare_seqtab.sh --help