- Version: 1.0
- Last modified: Thu Sep 3 12:52:30 CEST 2020
- Sign: Abhijeet Singh (abhijeetsingh.aau@gmail.com)
AcetoScan is a software pipeline for the analysis of Illumina MiSeq sequencing data for the FTHFS (Formyl--tetrahydrofolate synthetase) gene/amplicons
AcetoScan can also process fasta sequences to filter out non-target sequences, assigning taxonomy to the FTHFS fasta sequences and generate phylogenetic tree (see AcetoScan commands)
AcetoScan pipeline uses some software dependencies (version equals or higher)
- Cutadapt (2.9)
- VSEARCH (2.13.1)
- NCBI-blast+ (2.5.0+)
- Bioperl (1.7.2-3)
- MAFFT (7.307)
- Fasttree (2.1.9)
- R (3.5.2), with libraries:
¤ phyloseq (1.24.2)
¤ ggplot2 (3.1.1)
¤ plotly (4.9.0)
¤ RColorBrewer (1.1.2)
¤ plyr (1.8.4)
¤ dplyr (0.8.0.1)
¤ vegan (2.5.6)
For installation run the following command in terminal, this will check all dependencies and download the reference database from acetobase website.
$ chmod +x install_linux.sh OR install_mac.sh
$ sudo ./install_linux.sh OR install_mac.sh
For installation as local user make sure the dependency software are installed and modules are loaded (on server environment)
bash install_linux.sh OR install_mac.sh
install_linux.sh OR install_mac.sh will create main directory acetoscan in $HOME and sub-directories
1. bin - containing all main AcetoScan command scripts
2. dat - containing test
3. db - containing reference database AcetoBase
4. doc - containing user manual and tutorial video
5. scripts - containing dependencies scripts
1. user_data_directory - containing input raw data
https://support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/miseqreporter/miseq-reporter-generate-fastq-workflow-guide-15042322-01.pdf, page 9, FASTQ File Names
acetoscan will result in two directories
- Directories will be created
in default/destination path
1. output_data - containing process data will be generated and stored. In case of process failure, data can be accessed from here for further processing
2. acetoscan_result - containing all the final graphics, OTU table and TAX table. After successful execution of analysis, all the important data will be copies to this final directory.
$ acetoscan -X
or
$ /$HOME/acetoscan/acetoscan -X
1. acetoscan - for complete processing of raw sequence data
2. acetocheck - for processing fasta sequences and filter out non-target sequences
3. acetotax - acetocheck + taxonomic assignments
4. acetotree - acetotax + phylogenetic tree generation
Use acetoscan as follows
acetoscan -i /<input path>/ [-o /<output path>/] [-m 300] [-n 120] [-q 20] [-l 24] [-r 1] [-t 0.80] [-c 2] [-e 1e-3] [-B 1000] [-P 8]
-i Input directory containing raw illumina data
-o Output directory
:default = /$HOME/acetoscan/output_data
-m Maximum length of sequence after quality filtering
:default max_length = 300
-n Minimum length of sequence after quality filtering
:default min_length = 120
-q Quality threshold for the sequences
:default quality threshold = 20
-l Primer length
:default primer length = 24
-r Read type either forward or reverse reads
1 = forward reads (default), 2 = reverse reads
-t Clustering threshold
:default cluster threshold = 0.80 (80 %)
-c Minimum cluster size
:default minimum cluster size = 2
-e E-value
:default evalue = 1e-3
-B Bootstrap value
:default bootstrap = 1000
-P Parallel processes / threads
:default no. of parallels = all available threads
-h Print help
-X Print AcetoScan commands
-v Print AcetoScan version
-C Print AcetoScan citation
Use this command for the FTHFS fasta a sequences and filter out any unspecific / non-FTHFS sequence
$ acetocheck -h
acetocheck -i /path/<input_file> [-o /path/<output_file>] [-e 1e-3] [-P 8]
-i Input file - multifasta file
-o Output file
:default = acetocheck_<date>_<time>.fasta
-e E-value
:default evalue = 1e-3
-P Parallel processes/threads
:default no. of parallels = all available threads
-h Print help
-X Print AcetoScan commands
-v Print AcetoScan version
-C Print AcetoScan citation
acetocheck -i /home/abhi/Desktop/seq.fasta -o /home/abhi/Desktop/my_sequences -e 1e-3 -P 8
- my_sequences.fasta
- acetotax_< Date >_< Time >.log
Use this command for the FTHFS fasta a sequences and filter out any unspecific / non-FTHFS sequence and taxonomic annotations of the fasta sequences
$ acetotax -h
acetotax -i /path/<input_file>/ [-o /path/<output_file>/] [-e 1e-3] [-P 8]
-i Input_file
-o Output file
:default = acetotax_<date>_<time>.csv
:default = acetotax_<date>_<time>.fasta
-e E-value
:default evalue = 1e-3
-P Parallel processes/threads
:default no. of parallels = all available threads
-h Print help
-X Print AcetoScan commands
-v Print AcetoScan version
-C Print AcetoScan citation
acetotax -i /home/abhi/Desktop/seq.fasta -o /home/abhi/Desktop/my_sequences -e 1e-3 -P 8
- my_sequences.fasta
- my_sequences.csv
- acetotax_< Date >_< Time >.log
Use this command for the FTHFS fasta a sequences and filter out any unspecific / non-FTHFS sequence and taxonomic annotations of the fasta sequences and generation of phylogenetic tree
$ acetotree -h
acetotree -i /path/<input_file> [-o /path/<output_file>] [-e 1e-3] [-B 1000] [-P 8]
-i Input file - multifasta file
-o Output file
:default = acetotree_<date>_<time>.fasta
:default = acetotree_<date>_<time>.csv
:default = acetotree_<date>_<time>.aln
:default = acetotree_<date>_<time>.tree
-e E-value
:default evalue = 1e-3
-B Bootstrap value
:default bootstrap = 1000
-P Parallel processes/threads
:default no. of parallels = all available threads
-h Print help
-X Print AcetoScan commands
-v Print AcetoScan version
-C Print AcetoScan citation
acetotree -i /home/abhi/Desktop/seq.fasta -o /home/abhi/Desktop/my_sequences -e 1e-3 -B 1000 -P 8
- my_sequences.fasta
- my_sequences.csv
- my_sequences.aln
- my_sequences.tree
- acetotax_< Date >_< Time >.log
sudo docker volume create --opt type=none --opt o=bind --opt device=/PATH/to/my/DATA --name MY_CUSTOM_NAME
--opt device= - will have the path to your raw input data
--name - any name according to your wish
sudo docker volume create --opt type=none --opt o=bind --opt device=/home/abhi/Desktop/reads --name myDockerAcetoscan
sudo docker run --rm -v MY_CUSTOM_NAME:/acetoscan/input_dir -it abhijeetsingh1704/acetoscan -i /acetoscan/input_dir
sudo docker run --rm -v myDockerAcetoscan:/acetoscan/input_dir -it abhijeetsingh1704/acetoscan -i /acetoscan/input_dir
Default program for AcetoScan pipeline is acetoscan command, therefore it is optional to call it with --entrypoint flag. But in case of acetocheck,acetotax or acetotree command the code need to include --entrypoint flag
sudo docker run --rm -v myDockerAcetoscan:/acetoscan/input_dir --entrypoint acetoscan -it abhijeetsingh1704/acetoscan -i /acetoscan/input_dir
sudo docker run --rm -v myDockerAcetoscan:/acetoscan/input_dir --entrypoint acetocheck -it abhijeetsingh1704/acetoscan -i /acetoscan/input_dir/input_file.fasta
- OR
sudo docker run --rm -v myDockerAcetoscan:/acetoscan/input_dir --entrypoint acetocheck -it abhijeetsingh1704/acetoscan -i /acetoscan/input_dir/input_file.fasta -o /acetoscan/input_dir/output_file.fasta
Singh A, Nylander JAA, Schnürer A, Bongcam-Rudloff E and Müller B (2020) High-Throughput Sequencing and Unsupervised Analysis of Formyltetrahydrofolate Synthetase (FTHFS) Gene Amplicons to Estimate Acetogenic Community Structure. Front. Microbiol. 11:2066. doi: 10.3389/fmicb.2020.02066