A command-line tool to convert rpSBML files into SBOL and CSV files ready-to-be used with DNA-Bot.
rpbasicdesign extracts enzyme IDs from rpSBML files -- produced by the RP suite available in the SynBioCAD Galaxy platform -- to generate genetic constructs compliant with the BASIC assembly approach. CSV files produced are ready to be used with DNA-Bot to generate instructions for automated build of the genetic constructs using OpenTrons liquid handling robots.
conda install -c brsynth -c conda-forge rpbasicdesignSimple call:
conda activate <myenv>
python -m rpbasicdesign.cli --rpsbml_file tests/input/muconate_example.xmlOutput folders for dnabot-ready files and SBOL export can be set using o_dnabot_dir and o_sbol_dir options:
python -m rpbasicdesign.cli \
--rpsbml_file tests/input/muconate_example.xml \
--o_dnabot_dir out/dnabot_input \
--o_sbol_dir out/sbol_exportThe number of constructs to design is tuned using sample_size:
python -m rpbasicdesign.cli \
--rpsbml_file tests/input/muconate_example.xml \
--sample_size 5The complete list options is provided the embedded help, which can be printed using the --help or -h keywords:
python -m rpbasicdesign.cli -h
usage: python -m rpbasicdesign.cli [-h]
--rpsbml_file RPSBML_FILE
[--parts_files PARTS_FILES [PARTS_FILES ...]]
[--lms_id LMS_ID]
[--lmp_id LMP_ID]
[--backbone_id BACKBONE_ID]
[--sample_size SAMPLE_SIZE]
[--cds_permutation CDS_PERMUTATION]
[--max_enz_per_rxn MAX_ENZ_PER_RXN]
[--o_dnabot_dir O_DNABOT_DIR]
[--o_sbol_dir O_SBOL_DIR]
Convert rpSBML enzyme info in to BASIC construct. UniProt IDs corresponding enzyme variants are extracted rpSBMl files. Promoters and RBSs are randomly chosen from a default list. CDSs, in other words gene variants,
of enzymes are randomly chosen from amongst the UniProt IDs extracted. Constructs generated can be stored as (i) a CSV file ready to be used by DNA-Bot, (ii) as SBOL files.
optional arguments:
-h, --help show this help message and exit
--rpsbml_file RPSBML_FILE
rpSBML file from which enzymes UniProt IDs will be collected.
--parts_files PARTS_FILES [PARTS_FILES ...]
List of files providing available linkers and user parts (backbone, promoters, ...) for constructs. Default: [data/biolegio_parts.csv, user_parts.csv]
--lms_id LMS_ID part ID to be used as the LMS methylated linker. Default: LMS
--lmp_id LMP_ID part ID to be used as the LMP methylated linker. Default: LMP
--backbone_id BACKBONE_ID
part ID to be used as the backbone. Default: BASIC_SEVA_37_CmR-p15A.1
--sample_size SAMPLE_SIZE
Number of construct to generate.Default: 88
--cds_permutation CDS_PERMUTATION
Whether all combinations of CDS permutation should be built Default: true
--max_enz_per_rxn MAX_ENZ_PER_RXN
Maximum number of enyzme to consider per reaction. If more enzymes are available for a given reaction, then only the last one listed in the MIRIAM annotation section will be kept.
--max_gene_per_construct MAX_GENE_PER_CONSTRUCT
Maximum number of genes per construct. If more genes are required, i.e. more reactions are described in the inputet SBML file, then the execution will failed.
--o_dnabot_dir O_DNABOT_DIR
Output folder to write construct and plate files. It will be created if it does not exist yet. Existing files will be overwritten. Default: out/dnabot_in
--o_sbol_dir O_SBOL_DIR
Output folder to write SBOL depictions of constructs. Existing files will be overwritten. Default: not output.
If one wishes to only use a subset of BASIC parts, the way to go is to
provide a restricted list of parts with the --parts_file option.
The command below generates up to 88 constructs for the lycopene producing
pathway (CrtEBI pathway) defined in examples/lycopene_CrtEBI_from_selenzy.xml.xml, using the parts
described in examples/parts_for_lycopene.csv. Output files will be written
in examples/lycopene_sbol folder for SBOL files and examples/lycopene_dnabot
for DNA-Bot. At the end 88 constructs should be outputted.
python -m rpbasicdesign.cli --rpsbml_file examples/lycopene_CrtEBI_from_selenzy.xml --sample_size 88 --parts_files examples/parts_for_lycopene.csv --o_sbol_dir examples/lycopene_sbol_crtEBI --o_dnabot_dir examples/lycopene_dnabot_crtEBI --max_enz_per_rxn 1This section documents input files required / optional, their purpose, and how information should be structured.
SBML with retropath-like annotations. UnitProt IDs of enzyme are expected to be listed here. More information of rpSBML file at https://github.com/brsynth/rptools. Some examples or rpSBML files are provided in tests/input.
These are CSV files listing the linker IDs available for the constructs (BASIC linkers), as well as the user parts (backbone, promoters, ...). The format should be comma separated on 4 columns with header. Example below:
id,type,sequence,comment
L1,neutral linker,,
L2,neutral linker,,
L3,neutral linker,,
By default, the rpbasicdesign/data/biolegio_parts.csv file is used which corresponds to the BioLegio commercial plate (link). A second predefined file corresponding to older version of the BioLegio plate is also described in rpbasicdesign/data/legacy_parts.csv.
For linkers, the type annotation should be one of neutral linker, methylated linker, peptide fusion linker or RBS linker. For user parts, type should be one of backbone or constitutive promoter. Other type will raise a warning and will be omited. By default, biolegio_parts.csv and user_parts.csv are used.
Use the parts_files arguments to override.
Important:
- IDs should match the linker naming conventions (see below).
- IDs should match the IDs used in the plate file inputed to dnabot. As example -- but also ready to be used -- the biolegio_plate.csv is a valid input files for dnabot, with consistent IDs between
biolegio_parts.csvandbiolegio_plate.csv.
git clone https://github.com/brsynth/rpbasicdesign.git
cd rpbasicdesign
conda env create -f environment.yaml -n <myenv>
conda develop -n <myenv> .conda activate <myenv>
python -m pytest -v --cov=rpbasicdesign --cov-report htmlThe BASIC linker set is a major piece of the BASIC assembly method. For a detailed explanation of the BASIS approach, see Storch et. al., ACS Synth. Biol., 2015 (doi: 10.1021/sb500356d).
Only polycistronic constructs are enabled at the moment.
By default, the set of linkers used is the one presented available in from the commercial plate from BioLegio. If one wants to use its own set of linkers, the user is advised to do it carefully and to look for more information.
Due to DNA-Bot implementation:
- RBS linkers should start with the
Unsuffix, wherencould be any alphanumeric character. - Any linkers should have its two half linkers ending with the
-Pand-Ssuffixes listed in the "plate" file, ie in the file that provides the well locations containing the DNA fragment. See the BASIC approach paper, and especially the supplementary files for more information.
Parts and linkers provided in the *_parts.csv files have to match on the following type:
neutral linkermethylated linkerRBS linkerpeptide fusion linkerbackboneconstitutive promoter
As of today, CDS are obtained only by parsing rpSBML files.
For advanced users wishing to play with custom linkers:
- Linkers and parts can be provided using a custom file with the
--parts_filesargument. - Linkers described
user_parts.csvare not considered. - RBS linker IDs have to be in the form
AAA-BBBwithAAAbeing the linker suffix ID. - Linker prefixes and suffixes coordinates on the plate have to be listed in
[biolegio|legacy]_plate.csv.
The maximum number of genes in a construct limited to 3 with the default biolegio_plate.csv RBS library, because
there is only 3 different RBS suffix in the commercial BioLegio library. Anyway, if needed, this max number of genes
can be relaxed and increased using the --max_gene_per_construct parameter.
- Better handle logs and add
verboseoption
- Galaxy-SynBioCAD: https://doi.org/10.1101/2020.06.14.145730
- DNA-Bot: https://doi.org/10.1093/synbio/ysaa010
- BASIC assembly method: https://doi.org/10.1021/sb500356d