Source 1primary paper2021Genetics
Toolkit/plasmid toolkit for CeLINC
plasmid toolkit for CeLINC
Taxonomy: Mechanism Branch / Architecture. Workflows sit above the mechanism and technique branches rather than replacing them.
Summary
The plasmid toolkit for CeLINC is a construct set that extends the CeLINC optical binary protein-protein interaction assay in Caenorhabditis elegans by enabling custom promoter usage or substitution of CRY2 variants. It is intended to increase flexibility of a CRY2/CIB1-based in vivo colocalization assay for testing whether two proteins interact in vivo.
Usefulness & Problems
Why this is useful
This toolkit is useful because it broadens how CeLINC constructs can be configured, specifically by allowing custom promoters and alternative CRY2 variants. That flexibility can support adaptation of the in vivo interaction assay to different expression contexts within C. elegans.
Source:
We confirmed interactions seen between PAR-6, PKC-3, and PAR-3
Source:
but observed no physical interactions among the basolateral Scribble module proteins LET-413, DLG-1, and LGL-1.
Source:
We present Caenorhabditis elegans light-induced coclustering (CeLINC), an optical binary protein-protein interaction assay to determine whether two proteins interact in vivo.
Problem solved
It addresses the limited configurability of the original CeLINC construct system by providing plasmids that permit promoter replacement and CRY2 variant substitution. This helps tailor an optical protein-protein interaction assay to specific experimental expression requirements in C. elegans.
Source:
We confirmed interactions seen between PAR-6, PKC-3, and PAR-3
Source:
but observed no physical interactions among the basolateral Scribble module proteins LET-413, DLG-1, and LGL-1.
Taxonomy & Function
Primary hierarchy
Mechanism Branch
Architecture: A reusable architecture pattern for arranging parts into an engineered system.
Techniques
Functional AssayTarget processes
No target processes tagged yet.
Implementation Constraints
cofactor dependency: cofactor requirement unknownencoding mode: genetically encodedimplementation constraint: context specific validationoperating role: sensor
The toolkit is described as plasmid-based and compatible with the CeLINC assay in C. elegans. Available implementation details from the evidence are limited to support for custom promoters and CRY2 variant substitution; no further construct design, delivery, or expression details are provided here.
The evidence provided for the toolkit itself is limited to the statement that it enables custom promoters or CRY2 variants; no specific plasmid architectures, performance comparisons, or validation data for the modified constructs are given here. Independent replication and broad benchmarking of the toolkit are not documented in the supplied evidence.
Validation
Cell-freeBacteriaMammalianMouseHumanTherapeuticIndep. Replication
Supporting Sources
Ranked Claims
Using CeLINC, the authors confirmed interactions among the apical polarity regulators PAR-6, PKC-3, and PAR-3.
We confirmed interactions seen between PAR-6, PKC-3, and PAR-3
Using CeLINC, the authors confirmed interactions among the apical polarity regulators PAR-6, PKC-3, and PAR-3.
We confirmed interactions seen between PAR-6, PKC-3, and PAR-3
Using CeLINC, the authors confirmed interactions among the apical polarity regulators PAR-6, PKC-3, and PAR-3.
We confirmed interactions seen between PAR-6, PKC-3, and PAR-3
Using CeLINC, the authors confirmed interactions among the apical polarity regulators PAR-6, PKC-3, and PAR-3.
We confirmed interactions seen between PAR-6, PKC-3, and PAR-3
Using CeLINC, the authors confirmed interactions among the apical polarity regulators PAR-6, PKC-3, and PAR-3.
We confirmed interactions seen between PAR-6, PKC-3, and PAR-3
Using CeLINC, the authors confirmed interactions among the apical polarity regulators PAR-6, PKC-3, and PAR-3.
We confirmed interactions seen between PAR-6, PKC-3, and PAR-3
Using CeLINC, the authors confirmed interactions among the apical polarity regulators PAR-6, PKC-3, and PAR-3.
We confirmed interactions seen between PAR-6, PKC-3, and PAR-3
Using CeLINC, the authors observed no physical interactions among the basolateral Scribble module proteins LET-413, DLG-1, and LGL-1.
but observed no physical interactions among the basolateral Scribble module proteins LET-413, DLG-1, and LGL-1.
Using CeLINC, the authors observed no physical interactions among the basolateral Scribble module proteins LET-413, DLG-1, and LGL-1.
but observed no physical interactions among the basolateral Scribble module proteins LET-413, DLG-1, and LGL-1.
Using CeLINC, the authors observed no physical interactions among the basolateral Scribble module proteins LET-413, DLG-1, and LGL-1.
but observed no physical interactions among the basolateral Scribble module proteins LET-413, DLG-1, and LGL-1.
Using CeLINC, the authors observed no physical interactions among the basolateral Scribble module proteins LET-413, DLG-1, and LGL-1.
but observed no physical interactions among the basolateral Scribble module proteins LET-413, DLG-1, and LGL-1.
Using CeLINC, the authors observed no physical interactions among the basolateral Scribble module proteins LET-413, DLG-1, and LGL-1.
but observed no physical interactions among the basolateral Scribble module proteins LET-413, DLG-1, and LGL-1.
Using CeLINC, the authors observed no physical interactions among the basolateral Scribble module proteins LET-413, DLG-1, and LGL-1.
but observed no physical interactions among the basolateral Scribble module proteins LET-413, DLG-1, and LGL-1.
Using CeLINC, the authors observed no physical interactions among the basolateral Scribble module proteins LET-413, DLG-1, and LGL-1.
but observed no physical interactions among the basolateral Scribble module proteins LET-413, DLG-1, and LGL-1.
CeLINC is an optical binary protein-protein interaction assay for determining whether two proteins interact in vivo in Caenorhabditis elegans.
We present Caenorhabditis elegans light-induced coclustering (CeLINC), an optical binary protein-protein interaction assay to determine whether two proteins interact in vivo.
CeLINC is an optical binary protein-protein interaction assay for determining whether two proteins interact in vivo in Caenorhabditis elegans.
We present Caenorhabditis elegans light-induced coclustering (CeLINC), an optical binary protein-protein interaction assay to determine whether two proteins interact in vivo.
CeLINC is an optical binary protein-protein interaction assay for determining whether two proteins interact in vivo in Caenorhabditis elegans.
We present Caenorhabditis elegans light-induced coclustering (CeLINC), an optical binary protein-protein interaction assay to determine whether two proteins interact in vivo.
CeLINC is an optical binary protein-protein interaction assay for determining whether two proteins interact in vivo in Caenorhabditis elegans.
We present Caenorhabditis elegans light-induced coclustering (CeLINC), an optical binary protein-protein interaction assay to determine whether two proteins interact in vivo.
CeLINC is an optical binary protein-protein interaction assay for determining whether two proteins interact in vivo in Caenorhabditis elegans.
We present Caenorhabditis elegans light-induced coclustering (CeLINC), an optical binary protein-protein interaction assay to determine whether two proteins interact in vivo.
CeLINC is an optical binary protein-protein interaction assay for determining whether two proteins interact in vivo in Caenorhabditis elegans.
We present Caenorhabditis elegans light-induced coclustering (CeLINC), an optical binary protein-protein interaction assay to determine whether two proteins interact in vivo.
CeLINC is an optical binary protein-protein interaction assay for determining whether two proteins interact in vivo in Caenorhabditis elegans.
We present Caenorhabditis elegans light-induced coclustering (CeLINC), an optical binary protein-protein interaction assay to determine whether two proteins interact in vivo.
CeLINC is based on CRY2/CIB1 light-dependent oligomerization and detects interaction by colocalization of a fluorescently tagged prey protein with clustered bait protein.
Based on CRY2/CIB1 light-dependent oligomerization, CeLINC can rapidly and unambiguously identify protein-protein interactions between pairs of fluorescently tagged proteins. A fluorescently tagged bait protein is captured using a nanobody directed against the fluorescent protein (GFP or mCherry) and brought into artificial clusters within the cell. Colocalization of a fluorescently tagged prey protein in the cluster indicates a protein interaction.
CeLINC is based on CRY2/CIB1 light-dependent oligomerization and detects interaction by colocalization of a fluorescently tagged prey protein with clustered bait protein.
Based on CRY2/CIB1 light-dependent oligomerization, CeLINC can rapidly and unambiguously identify protein-protein interactions between pairs of fluorescently tagged proteins. A fluorescently tagged bait protein is captured using a nanobody directed against the fluorescent protein (GFP or mCherry) and brought into artificial clusters within the cell. Colocalization of a fluorescently tagged prey protein in the cluster indicates a protein interaction.
CeLINC is based on CRY2/CIB1 light-dependent oligomerization and detects interaction by colocalization of a fluorescently tagged prey protein with clustered bait protein.
Based on CRY2/CIB1 light-dependent oligomerization, CeLINC can rapidly and unambiguously identify protein-protein interactions between pairs of fluorescently tagged proteins. A fluorescently tagged bait protein is captured using a nanobody directed against the fluorescent protein (GFP or mCherry) and brought into artificial clusters within the cell. Colocalization of a fluorescently tagged prey protein in the cluster indicates a protein interaction.
CeLINC is based on CRY2/CIB1 light-dependent oligomerization and detects interaction by colocalization of a fluorescently tagged prey protein with clustered bait protein.
Based on CRY2/CIB1 light-dependent oligomerization, CeLINC can rapidly and unambiguously identify protein-protein interactions between pairs of fluorescently tagged proteins. A fluorescently tagged bait protein is captured using a nanobody directed against the fluorescent protein (GFP or mCherry) and brought into artificial clusters within the cell. Colocalization of a fluorescently tagged prey protein in the cluster indicates a protein interaction.
CeLINC is based on CRY2/CIB1 light-dependent oligomerization and detects interaction by colocalization of a fluorescently tagged prey protein with clustered bait protein.
Based on CRY2/CIB1 light-dependent oligomerization, CeLINC can rapidly and unambiguously identify protein-protein interactions between pairs of fluorescently tagged proteins. A fluorescently tagged bait protein is captured using a nanobody directed against the fluorescent protein (GFP or mCherry) and brought into artificial clusters within the cell. Colocalization of a fluorescently tagged prey protein in the cluster indicates a protein interaction.
CeLINC is based on CRY2/CIB1 light-dependent oligomerization and detects interaction by colocalization of a fluorescently tagged prey protein with clustered bait protein.
Based on CRY2/CIB1 light-dependent oligomerization, CeLINC can rapidly and unambiguously identify protein-protein interactions between pairs of fluorescently tagged proteins. A fluorescently tagged bait protein is captured using a nanobody directed against the fluorescent protein (GFP or mCherry) and brought into artificial clusters within the cell. Colocalization of a fluorescently tagged prey protein in the cluster indicates a protein interaction.
CeLINC is based on CRY2/CIB1 light-dependent oligomerization and detects interaction by colocalization of a fluorescently tagged prey protein with clustered bait protein.
Based on CRY2/CIB1 light-dependent oligomerization, CeLINC can rapidly and unambiguously identify protein-protein interactions between pairs of fluorescently tagged proteins. A fluorescently tagged bait protein is captured using a nanobody directed against the fluorescent protein (GFP or mCherry) and brought into artificial clusters within the cell. Colocalization of a fluorescently tagged prey protein in the cluster indicates a protein interaction.
In the reported reference-pair testing, CeLINC showed extremely robust assay performance with no false positives detected in the negative reference pairs.
We tested the system with an array of positive and negative reference protein pairs. Assay performance was extremely robust with no false positives detected in the negative reference pairs.
In the reported reference-pair testing, CeLINC showed extremely robust assay performance with no false positives detected in the negative reference pairs.
We tested the system with an array of positive and negative reference protein pairs. Assay performance was extremely robust with no false positives detected in the negative reference pairs.
In the reported reference-pair testing, CeLINC showed extremely robust assay performance with no false positives detected in the negative reference pairs.
We tested the system with an array of positive and negative reference protein pairs. Assay performance was extremely robust with no false positives detected in the negative reference pairs.
In the reported reference-pair testing, CeLINC showed extremely robust assay performance with no false positives detected in the negative reference pairs.
We tested the system with an array of positive and negative reference protein pairs. Assay performance was extremely robust with no false positives detected in the negative reference pairs.
In the reported reference-pair testing, CeLINC showed extremely robust assay performance with no false positives detected in the negative reference pairs.
We tested the system with an array of positive and negative reference protein pairs. Assay performance was extremely robust with no false positives detected in the negative reference pairs.
In the reported reference-pair testing, CeLINC showed extremely robust assay performance with no false positives detected in the negative reference pairs.
We tested the system with an array of positive and negative reference protein pairs. Assay performance was extremely robust with no false positives detected in the negative reference pairs.
In the reported reference-pair testing, CeLINC showed extremely robust assay performance with no false positives detected in the negative reference pairs.
We tested the system with an array of positive and negative reference protein pairs. Assay performance was extremely robust with no false positives detected in the negative reference pairs.
The CeLINC plasmid toolkit allows use of custom promoters or CRY2 variants and supports use with existing fluorescently tagged strains without additional cloning or genome modification.
We have generated a plasmid toolkit that allows use of custom promoters or CRY2 variants to promote flexibility of the system. The CeLINC assay is a powerful and rapid technique that can be widely applied in C. elegans due to the universal plasmids that can be used with existing fluorescently tagged strains without need for additional cloning or genetic modification of the genome.
The CeLINC plasmid toolkit allows use of custom promoters or CRY2 variants and supports use with existing fluorescently tagged strains without additional cloning or genome modification.
We have generated a plasmid toolkit that allows use of custom promoters or CRY2 variants to promote flexibility of the system. The CeLINC assay is a powerful and rapid technique that can be widely applied in C. elegans due to the universal plasmids that can be used with existing fluorescently tagged strains without need for additional cloning or genetic modification of the genome.
The CeLINC plasmid toolkit allows use of custom promoters or CRY2 variants and supports use with existing fluorescently tagged strains without additional cloning or genome modification.
We have generated a plasmid toolkit that allows use of custom promoters or CRY2 variants to promote flexibility of the system. The CeLINC assay is a powerful and rapid technique that can be widely applied in C. elegans due to the universal plasmids that can be used with existing fluorescently tagged strains without need for additional cloning or genetic modification of the genome.
The CeLINC plasmid toolkit allows use of custom promoters or CRY2 variants and supports use with existing fluorescently tagged strains without additional cloning or genome modification.
We have generated a plasmid toolkit that allows use of custom promoters or CRY2 variants to promote flexibility of the system. The CeLINC assay is a powerful and rapid technique that can be widely applied in C. elegans due to the universal plasmids that can be used with existing fluorescently tagged strains without need for additional cloning or genetic modification of the genome.
The CeLINC plasmid toolkit allows use of custom promoters or CRY2 variants and supports use with existing fluorescently tagged strains without additional cloning or genome modification.
We have generated a plasmid toolkit that allows use of custom promoters or CRY2 variants to promote flexibility of the system. The CeLINC assay is a powerful and rapid technique that can be widely applied in C. elegans due to the universal plasmids that can be used with existing fluorescently tagged strains without need for additional cloning or genetic modification of the genome.
The CeLINC plasmid toolkit allows use of custom promoters or CRY2 variants and supports use with existing fluorescently tagged strains without additional cloning or genome modification.
We have generated a plasmid toolkit that allows use of custom promoters or CRY2 variants to promote flexibility of the system. The CeLINC assay is a powerful and rapid technique that can be widely applied in C. elegans due to the universal plasmids that can be used with existing fluorescently tagged strains without need for additional cloning or genetic modification of the genome.
The CeLINC plasmid toolkit allows use of custom promoters or CRY2 variants and supports use with existing fluorescently tagged strains without additional cloning or genome modification.
We have generated a plasmid toolkit that allows use of custom promoters or CRY2 variants to promote flexibility of the system. The CeLINC assay is a powerful and rapid technique that can be widely applied in C. elegans due to the universal plasmids that can be used with existing fluorescently tagged strains without need for additional cloning or genetic modification of the genome.
Approval Evidence
1 source1 linked approval claimfirst-pass slug plasmid-toolkit-for-celinc
We have generated a plasmid toolkit that allows use of custom promoters or CRY2 variants to promote flexibility of the system.
Source:
usabilitysupports
The CeLINC plasmid toolkit allows use of custom promoters or CRY2 variants and supports use with existing fluorescently tagged strains without additional cloning or genome modification.
We have generated a plasmid toolkit that allows use of custom promoters or CRY2 variants to promote flexibility of the system. The CeLINC assay is a powerful and rapid technique that can be widely applied in C. elegans due to the universal plasmids that can be used with existing fluorescently tagged strains without need for additional cloning or genetic modification of the genome.
Source:
Comparisons
Source-backed strengths
The main strength supported by the evidence is increased system flexibility through custom promoters or CRY2 variants. The underlying CeLINC assay was applied to confirm interactions among PAR-6, PKC-3, and PAR-3 and to observe no physical interactions among LET-413, DLG-1, and LGL-1, indicating that the parent assay can distinguish positive and negative interaction cases in vivo.
Source:
We tested the system with an array of positive and negative reference protein pairs. Assay performance was extremely robust with no false positives detected in the negative reference pairs.
Compared with CheRiff
plasmid toolkit for CeLINC and CheRiff address a similar problem space.
Shared frame: same top-level item type
Strengths here: looks easier to implement in practice.
Compared with intermolecular disulfide-based light switch
plasmid toolkit for CeLINC and intermolecular disulfide-based light switch address a similar problem space.
Shared frame: same top-level item type
Strengths here: looks easier to implement in practice.
plasmid toolkit for CeLINC and Pyr-NHS-functionalised 3D graphene foam electrode biosensor address a similar problem space.
Shared frame: same top-level item type
Ranked Citations
- 1.