MoClo Kit

An implementation of the original MoClo ToolKit for the Python MoClo library.

https://media.springernature.com/original/springer-static/image/chp%3A10.1007%2F978-1-0716-0908-8_8/MediaObjects/468902_1_En_8_Fig2_HTML.png

References

  1. Weber, E., Engler, C., Gruetzner, R., Werner, S., Marillonnet, S. (2011). A Modular Cloning System for Standardized Assembly of Multigene Constructs. PLOS ONE, 6(2), e16765..

  2. Werner, S., Engler, C., Weber, E., Gruetzner, R., & Marillonnet, S. (2012). Fast track assembly of multigene constructs using Golden Gate cloning and the MoClo system. Bioengineered, 3(1), 38–43..

  3. Marillonnet, S., Werner, S. (2020). Assembly of Multigene Constructs Using the Modular Cloning System MoClo. Methods Mol Biol. 2020;2205:125-141..

Level -1

Module

class moclo.kits.moclo.MoCloProduct(Product)[source]

An original MoClo product.

__init__(record)
cutter

alias of Bio.Restriction.Restriction.BpiI

is_valid()

Check if the wrapped record follows the required class structure.

Returns

True if the record is valid, False otherwise.

Return type

bool

overhang_end()

Get the downstream overhang of the target sequence.

Returns

the downstream overhang.

Return type

Seq

overhang_start()

Get the upstream overhang of the target sequence.

Returns

the downstream overhang.

Return type

Seq

classmethod structure()

Get the module structure, as a DNA regex pattern.

Warning

If overloading this method, the returned pattern must include 3 capture groups to capture the following features:

  1. The upstream (5’) overhang sequence

  2. The module target sequence

  3. The downstream (3’) overhang sequence

target_sequence()

Get the target sequence of the module.

Modules are often stored in a standardized way, and contain more than the sequence of interest: for instance they can contain an antibiotic marker, that will not be part of the assembly when that module is assembled into a vector; only the target sequence is inserted.

Returns

the target sequence with annotations.

Return type

SeqRecord

Note

Depending on the cutting direction of the restriction enzyme used during assembly, the overhang will be left at the beginning or at the end, so the obtained record is exactly the sequence the enzyme created during restriction.

Vector

class moclo.kits.moclo.MoCloEntryVector(EntryVector)[source]

A MoClo entry vector.

References

Weber et al., Figure 2A.

__init__(record)
assemble(module, *modules, **kwargs)

Assemble the provided modules into the vector.

Parameters

  • module (AbstractModule) – a module to insert in the vector.

  • modules (AbstractModule, optional) – additional modules to insert in the vector. The order of the parameters is not important, since modules will be sorted by their start overhang in the function.

Returns

the assembled sequence with sequence annotations inherited from the vector and the modules.

Return type

SeqRecord

Raises

  • DuplicateModules – when two different modules share the same start overhang, leading in possibly non-deterministic constructs.

  • MissingModule – when a module has an end overhang that is not shared by any other module, leading to a partial construct only

  • InvalidSequence – when one of the modules does not match the required module structure (missing site, wrong overhang, etc.).

  • UnusedModules – when some modules were not used during the assembly (mostly caused by duplicate parts).

cutter

alias of Bio.Restriction.Restriction.BpiI

is_valid()

Check if the wrapped record follows the required class structure.

Returns

True if the record is valid, False otherwise.

Return type

bool

overhang_end()

Get the downstream overhang of the vector sequence.

overhang_start()

Get the upstream overhang of the vector sequence.

placeholder_sequence()

Get the placeholder sequence in the vector.

The placeholder sequence is replaced by the concatenation of modules during the assembly. It often contains a dropout sequence, such as a GFP expression cassette that can be used to measure the progress of the assembly.

classmethod structure()[source]

Get the vector structure, as a DNA regex pattern.

Warning

If overloading this method, the returned pattern must include 3 capture groups to capture the following features:

  1. The downstream (3’) overhang sequence

  2. The vector placeholder sequence

  3. The upstream (5’) overhang sequence

target_sequence()

Get the target sequence in the vector.

The target sequence if the part of the plasmid that is not discarded during the assembly (everything except the placeholder sequence).

Level 0

Module

class moclo.kits.moclo.MoCloEntry(Entry)[source]

An original MoClo entry.

__init__(record)
cutter

alias of Bio.Restriction.Restriction.BsaI

is_valid()

Check if the wrapped record follows the required class structure.

Returns

True if the record is valid, False otherwise.

Return type

bool

overhang_end()

Get the downstream overhang of the target sequence.

Returns

the downstream overhang.

Return type

Seq

overhang_start()

Get the upstream overhang of the target sequence.

Returns

the downstream overhang.

Return type

Seq

classmethod structure()

Get the module structure, as a DNA regex pattern.

Warning

If overloading this method, the returned pattern must include 3 capture groups to capture the following features:

  1. The upstream (5’) overhang sequence

  2. The module target sequence

  3. The downstream (3’) overhang sequence

target_sequence()

Get the target sequence of the module.

Modules are often stored in a standardized way, and contain more than the sequence of interest: for instance they can contain an antibiotic marker, that will not be part of the assembly when that module is assembled into a vector; only the target sequence is inserted.

Returns

the target sequence with annotations.

Return type

SeqRecord

Note

Depending on the cutting direction of the restriction enzyme used during assembly, the overhang will be left at the beginning or at the end, so the obtained record is exactly the sequence the enzyme created during restriction.

Vector

class moclo.kits.moclo.MoCloCassetteVector(CassetteVector)[source]

A MoClo cassette vector.

References

Weber et al., Figure 4A.

__init__(record)
assemble(module, *modules, **kwargs)

Assemble the provided modules into the vector.

Parameters

  • module (AbstractModule) – a module to insert in the vector.

  • modules (AbstractModule, optional) – additional modules to insert in the vector. The order of the parameters is not important, since modules will be sorted by their start overhang in the function.

Returns

the assembled sequence with sequence annotations inherited from the vector and the modules.

Return type

SeqRecord

Raises

  • DuplicateModules – when two different modules share the same start overhang, leading in possibly non-deterministic constructs.

  • MissingModule – when a module has an end overhang that is not shared by any other module, leading to a partial construct only

  • InvalidSequence – when one of the modules does not match the required module structure (missing site, wrong overhang, etc.).

  • UnusedModules – when some modules were not used during the assembly (mostly caused by duplicate parts).

cutter

alias of Bio.Restriction.Restriction.BsaI

is_valid()

Check if the wrapped record follows the required class structure.

Returns

True if the record is valid, False otherwise.

Return type

bool

overhang_end()

Get the downstream overhang of the vector sequence.

overhang_start()

Get the upstream overhang of the vector sequence.

placeholder_sequence()

Get the placeholder sequence in the vector.

The placeholder sequence is replaced by the concatenation of modules during the assembly. It often contains a dropout sequence, such as a GFP expression cassette that can be used to measure the progress of the assembly.

classmethod structure()[source]

Get the vector structure, as a DNA regex pattern.

Warning

If overloading this method, the returned pattern must include 3 capture groups to capture the following features:

  1. The downstream (3’) overhang sequence

  2. The vector placeholder sequence

  3. The upstream (5’) overhang sequence

target_sequence()

Get the target sequence in the vector.

The target sequence if the part of the plasmid that is not discarded during the assembly (everything except the placeholder sequence).

Parts

class moclo.kits.moclo.MoCloPro(MoCloPart, MoCloEntry)[source]

An original MoClo promoter part.

class moclo.kits.moclo.MoClo5U(MoCloPart, MoCloEntry)[source]

An original MoClo 5’ UTR part.

class moclo.kits.moclo.MoClo5Uf(MoCloPart, MoCloEntry)[source]

An original MoClo 5’UTR part for N-terminal tag linkage.

class moclo.kits.moclo.MoCloNTag(MoCloPart, MoCloEntry)[source]

An original MoClo N-terminal tag part.

class moclo.kits.moclo.MoCloPro5U(MoCloPart, MoCloEntry)[source]

An original MoClo promoter fused with a 5’UTR part.

class moclo.kits.moclo.MoCloPro5Uf(MoCloPart, MoCloEntry)[source]

An original MoClo promoter fused with a 5’UTR for N-terminal linkage.

class moclo.kits.moclo.MoCloCDS1(MoCloPart, MoCloEntry)[source]

An original MoClo CDS1.

class moclo.kits.moclo.MoCloCDS1ns(MoCloPart, MoCloEntry)[source]

An original MoClo CDS1 without STOP codon for C-terminal tag linkage.

class moclo.kits.moclo.MoCloSP(MoCloPart, MoCloEntry)[source]

An original MoClo signal peptide part.

class moclo.kits.moclo.MoCloCDS2(MoCloPart, MoCloEntry)[source]

An original MoClo CDS2 part.

class moclo.kits.moclo.MoCloCDS2ns(MoCloPart, MoCloEntry)[source]

An original MoClo CDS2 for C-terminal tag linkage.

class moclo.kits.moclo.MoCloCTag(MoCloPart, MoCloEntry)[source]

An original MoClo C-terminal tag part.

class moclo.kits.moclo.MoClo3U(MoCloPart, MoCloEntry)[source]

An original MoClo 3’UTR part.

class moclo.kits.moclo.MoCloTer(MoCloPart, MoCloEntry)[source]

An original MoClo terminator part.

class moclo.kits.moclo.MoClo3UTer(MoCloPart, MoCloEntry)[source]

An original MoClo terminator part.

class moclo.kits.moclo.MoCloGene(MoCloPart, MoCloEntry)[source]

An complete transcription unit stored as an original MoClo part.

Level 1

Module

class moclo.kits.moclo.MoCloCassette(Cassette)[source]

An original MoClo cassette.

cutter

alias of Bio.Restriction.Restriction.BpiI

Vector

class moclo.kits.moclo.MoCloDeviceVector(DeviceVector)[source]

An original MoClo device vector.

References

Weber et al., Figure 4A.

cutter

alias of Bio.Restriction.Restriction.BpiI

Parts

class moclo.kits.moclo.MoCloEndLinker(MoCloPart, MoCloCassette)[source]

An Icon Genetic end linker part.

References

Weber et al., Figure 5.

Level M

Parts

class moclo.kits.moclo.MoCloLevelMVector(MoCloPart, MoCloDeviceVector)[source]
cutter

alias of Bio.Restriction.Restriction.BpiI

classmethod structure()[source]

Get the part structure, as a DNA regex pattern.

The structure of most parts can be obtained automatically from the part signature and the restriction enzyme used in the Golden Gate assembly.

Warning

If overloading this method, the returned pattern must include 3 capture groups to capture the following features:

  1. The upstream (5’) overhang sequence

  2. The vector placeholder sequence

  3. The downstream (3’) overhang sequence

class moclo.kits.moclo.MoCloLevelMEndLinker(MoCloPart, MoCloCassette)[source]
classmethod structure()[source]

Get the part structure, as a DNA regex pattern.

The structure of most parts can be obtained automatically from the part signature and the restriction enzyme used in the Golden Gate assembly.

Warning

If overloading this method, the returned pattern must include 3 capture groups to capture the following features:

  1. The upstream (5’) overhang sequence

  2. The vector placeholder sequence

  3. The downstream (3’) overhang sequence

Level P

Parts

class moclo.kits.moclo.MoCloLevelPVector(MoCloPart, MoCloCassetteVector)[source]
cutter

alias of Bio.Restriction.Restriction.BsaI

classmethod structure()[source]

Get the part structure, as a DNA regex pattern.

The structure of most parts can be obtained automatically from the part signature and the restriction enzyme used in the Golden Gate assembly.

Warning

If overloading this method, the returned pattern must include 3 capture groups to capture the following features:

  1. The upstream (5’) overhang sequence

  2. The vector placeholder sequence

  3. The downstream (3’) overhang sequence

class moclo.kits.moclo.MoCloLevelPEndLinker(MoCloPart, MoCloEntry)[source]
cutter

alias of Bio.Restriction.Restriction.BsaI

classmethod structure()[source]

Get the part structure, as a DNA regex pattern.

The structure of most parts can be obtained automatically from the part signature and the restriction enzyme used in the Golden Gate assembly.

Warning

If overloading this method, the returned pattern must include 3 capture groups to capture the following features:

  1. The upstream (5’) overhang sequence

  2. The vector placeholder sequence

  3. The downstream (3’) overhang sequence