Source 1primary paper2025MED
Toolkit/ribozyme-flanked gRNA
ribozyme-flanked gRNA
Also known as: RGR
Taxonomy: Mechanism Branch / Component. Workflows sit above the mechanism and technique branches rather than replacing them.
Summary
We engineered BLU-VIPR around... ribozyme-flanked gRNA. The BLU-VIPR design is genetically encoded and ensures precise excision of multiple gRNAs from a single messenger RNA transcript.
Usefulness & Problems
Why this is useful
The ribozyme-flanked gRNA design enables precise excision of guide RNAs from a single messenger RNA transcript. In BLU-VIPR this is the core mechanism for light-induced guide production.; precise excision of gRNAs from a single messenger RNA transcript; multiplex gRNA expression; genetically encoded gRNA production
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The ribozyme-flanked gRNA design enables precise excision of guide RNAs from a single messenger RNA transcript. In BLU-VIPR this is the core mechanism for light-induced guide production.
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precise excision of gRNAs from a single messenger RNA transcript
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multiplex gRNA expression
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genetically encoded gRNA production
Problem solved
It solves the problem of producing one or more functional gRNAs from a Pol II-like transcript under optogenetic transcriptional control. This supports multiplexed and simplified spatiotemporal CRISPR perturbation.; enables precise release of multiple gRNAs from one transcript
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It solves the problem of producing one or more functional gRNAs from a Pol II-like transcript under optogenetic transcriptional control. This supports multiplexed and simplified spatiotemporal CRISPR perturbation.
Source:
enables precise release of multiple gRNAs from one transcript
Problem links
enables precise release of multiple gRNAs from one transcript
LiteratureIt solves the problem of producing one or more functional gRNAs from a Pol II-like transcript under optogenetic transcriptional control. This supports multiplexed and simplified spatiotemporal CRISPR perturbation.
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It solves the problem of producing one or more functional gRNAs from a Pol II-like transcript under optogenetic transcriptional control. This supports multiplexed and simplified spatiotemporal CRISPR perturbation.
Published Workflows
Objective: Engineer an optogenetic CRISPR system that enables precise spatiotemporal gene perturbation by light-controlled guide RNA production, including in vivo use in T lymphocytes.
Why it works: The paper frames the approach as combining CRISPR flexibility with light's spatiotemporal resolution, while using a genetically encoded design that precisely excises multiple gRNAs from one transcript.
blue-light activated transcriptionoptogenetic regulation of gRNA productionribozyme-mediated excision of multiple gRNAs from a single transcriptengineering around a light-activated transcription factorengineering ribozyme-flanked gRNA expression architecture
Taxonomy & Function
Primary hierarchy
Mechanism Branch
Component: A low-level RNA part used inside a larger architecture that realizes a mechanism.
Techniques
Computational DesignTarget processes
transcriptionImplementation Constraints
cofactor dependency: cofactor requirement unknownencoding mode: genetically encodedimplementation constraint: context specific validationoperating role: regulator
It requires a genetically encoded transcript containing the ribozyme-flanked guide architecture. The abstract does not specify the exact ribozymes, although the web summary identifies HH and HDV in the anchor study.; must be embedded in a transcript architecture compatible with ribozyme-mediated excision
Independent follow-up evidence is still limited. Validation breadth across biological contexts is still narrow. Independent reuse still looks limited, so the evidence base may be fragile. No canonical validation observations are stored yet, so context-specific performance remains under-specified.
Validation
Cell-freeBacteriaMammalianMouseHumanTherapeuticIndep. Replication
Supporting Sources
Ranked Claims
BLU-VIPR enabled several types of optogenetic CRISPR including indels, CRISPRa, and base editing.
This simplified spatiotemporal gene perturbation and allowed for several types of optogenetic CRISPR, including indels, CRISPRa, and base editing.
BLU-VIPR was engineered around VPR-EL222 and ribozyme-flanked gRNA.
We engineered BLU-VIPR around a new potent blue-light activated transcription factor (VPR-EL222) and ribozyme-flanked gRNA.
The BLU-VIPR design ensures precise excision of multiple gRNAs from a single messenger RNA transcript.
The BLU-VIPR design is genetically encoded and ensures precise excision of multiple gRNAs from a single messenger RNA transcript.
BLU-VIPR worked in vivo and achieved optogenetic gene editing in T lymphocytes in vivo.
BLU-VIPR also worked in vivo with cells previously intractable to optogenetic gene editing, achieving optogenetic gene editing in T lymphocytes in vivo.
BLU-VIPR diverges from split-Cas strategies by optogenetically regulating gRNA production.
BLU-VIPR) that diverges from prevailing split-Cas design strategies and instead focuses on optogenetic regulation of guide RNA (gRNA) production.
BLU-VIPR is a new optogenetic CRISPR tool for light-controlled gene perturbation.
Here, we present a new optogenetic CRISPR tool (Blue Light-inducible Universal VPR-Improved Production of RGRs, BLU-VIPR)...
Approval Evidence
1 source2 linked approval claimsfirst-pass slug ribozyme-flanked-grna
We engineered BLU-VIPR around... ribozyme-flanked gRNA. The BLU-VIPR design is genetically encoded and ensures precise excision of multiple gRNAs from a single messenger RNA transcript.
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design featuresupports
BLU-VIPR was engineered around VPR-EL222 and ribozyme-flanked gRNA.
We engineered BLU-VIPR around a new potent blue-light activated transcription factor (VPR-EL222) and ribozyme-flanked gRNA.
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functional capabilitysupports
The BLU-VIPR design ensures precise excision of multiple gRNAs from a single messenger RNA transcript.
The BLU-VIPR design is genetically encoded and ensures precise excision of multiple gRNAs from a single messenger RNA transcript.
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Comparisons
Source-stated alternatives
The abstract does not name direct alternatives for guide processing. It only states that the overall system differs from split-Cas strategies.
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The abstract does not name direct alternatives for guide processing. It only states that the overall system differs from split-Cas strategies.
Source-backed strengths
ensures precise excision; supports multiple gRNAs from a single transcript; genetically encoded
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ensures precise excision
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supports multiple gRNAs from a single transcript
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genetically encoded
Compared with CIb1 promoter
ribozyme-flanked gRNA and CIb1 promoter address a similar problem space because they share transcription.
Shared frame: same top-level item type; shared target processes: transcription
Compared with microRNA
ribozyme-flanked gRNA and microRNA address a similar problem space because they share transcription.
Shared frame: same top-level item type; shared target processes: transcription
Strengths here: looks easier to implement in practice.
Compared with upstream ORFs
ribozyme-flanked gRNA and upstream ORFs address a similar problem space because they share transcription.
Shared frame: same top-level item type; shared target processes: transcription
Strengths here: looks easier to implement in practice.
Ranked Citations
- 1.