CRISPRi Gene Repression in Yeast

Precision Gene Silencing in Saccharomyces cerevisiae

Custom design and validation of dCas9/dCas12a-mediated gene repression for your target genes.

Custom gRNA design targeting promoter or ORF regions
Inducible or constitutive repression options
Validation by qRT-PCR or fluorescence-based readout

Service Overview

CRISPR interference (CRISPRi) enables targeted, reversible gene repression in Saccharomyces cerevisiae without introducing double-strand breaks. Our service provides end-to-end support from guide RNA design to validated engineered strains.

Target-specific gRNA libraries
DCas9 or dCas12a repressor systems
Repression efficiency data with QC metrics

Inducible Control

Multiplexing Options

Strain Validation

Our Approach

End-to-End CRISPRi Service for S. cerevisiae

We design, build, and validate CRISPRi constructs tailored to your project goals.

Custom gRNA Design & Cloning

Guide RNAs are designed using proprietary algorithms to evaluate on-target repression while minimizing off-target effects. gRNAs are cloned into expression vectors encoding a catalytically dead Cas9 or Cas12a fused to a transcriptional repressor domain.

Target gene sequence analysis
Optimal gRNA selection (promoter or ORF targeting)
Vector construction with dCas9-repressor and gRNA

Inducible Repression Control

Our system supports inducible CRISPRi using β-estradiol-responsive promoters, allowing temporal control of gene repression. Constitutive expression is also available when constant silencing is preferred.

Inducible (estradiol) or constitutive promoter options
Dose-dependent repression tuning
Suitable for essential gene studies and metabolic pathway modulation

Verified Delivery Vectors

We provide validated dCas9/dCas12a repressor constructs and gRNA expression cassettes for transformation into S. cerevisiae strains.

Robust QC Validation

Scoped per project after consultation.

Flexible Project Scope

Services are scaled to your needs—single gene targets, multiplexed repression, or library-scale screens—with deliverables defined per SOW.

Deliverables

What You Receive

Our standard handoff includes engineered strains, validation data, and project documentation.

Core Deliverables

Engineered S. cerevisiae strains with confirmed CRISPRi constructs
GRNA sequences and plasmid maps
Repression validation report (qRT-PCR or fluorescence data)

Additional Materials

QC data including growth curves and repression kinetics
Glycerol stocks of engineered strains
Project documentation with methods and results

Scope of Service

Customizable Parameters for Your Project

All aspects are defined during consultation to match your research needs.

Parameter	Typical project scope
Target Gene Selection	Single gene, gene family, or pathway; specified by customer or recommended by our bioinformatics team
GRNA Design & Synthesis	2-4 gRNAs per target; design based on promoter/ORF region; synthesis included
CRISPRi System	DCas9-Mxi1, dCas9-MIG1, or dCas12a-repressor; choice depends on PAM preference and target AT-content
Inducibility	β-estradiol inducible or constitutive expression; induction conditions optimized per strain
Multiplexing	Up to 4 gRNAs per vector (dCas12a array); multiple target repression in a single transformation
Strain Background	Laboratory strains (e.g., BY4741, W303) or industrial strains; compatibility assessed
Validation Method	QRT-PCR or fluorescence reporter (e.g., GFP) depending on target; fold-repression reported
Number of Clones	Typically 3–5 verified clones per target; more upon request
Technical Support	Named scientific contact at project start; milestone review calls; email response within 1 business day

Workflow

From Design to Validated Strain

A streamlined process with clear milestones and deliverables at each stage.

Guide RNA Design

Our bioinformatics team identifies optimal gRNA sequences targeting your gene(s) of interest. We evaluate promoter versus ORF targeting for maximal repression, and design multiple candidates for best efficiency.

Vector Construction

GRNA oligonucleotides are cloned into expression plasmids containing dCas9 or dCas12a fused to a repressor domain. The final construct is sequence-verified before transformation.

Yeast Transformation

The CRISPRi plasmid is transformed into your chosen S. cerevisiae strain using a standard lithium acetate protocol. Successful transformants are selected on appropriate auxotrophic or antibiotic markers.

Induction & Selection

For inducible systems, repression is triggered by adding β-estradiol to the culture medium. Cells are grown under selective pressure to maintain the plasmid, and repression is allowed to establish over several generations.

Validation & Handoff

Repression efficiency is quantified by qRT-PCR or fluorescence reading. At least three independent clones per target are characterized. Data and engineered strains are delivered in a final report.

Applications

Where CRISPRi in Yeast Is Used

Our service supports a wide range of research and biotechnology needs.

Pathway Flux Redirection

Fine-tune expression of endogenous or heterologous genes to redirect carbon flux toward desired products.

Repress competing metabolic branches
Balance cofactor regeneration
Modulate enzyme expression levels

Gene Essentiality & Phenotype Screens

Identify genes required for growth under specific conditions or uncover new functions in uncharacterized open reading frames.

Pooled library screening for fitness
Conditional essential gene studies
Target validation for drug discovery

Improving Industrial Robustness

Identify and repress negative regulators of stress tolerance to evaluate performance in industrial fermentation environments.

Acetic acid, ethanol, or osmotic stress
Temperature tolerance
Inhibitor resistance in lignocellulosic hydrolysates

FAQ

Frequently Asked Questions

Common questions about our CRISPRi gene repression service.

Repression efficiency is target- and locus-dependent. In typical projects, we observe project-dependent performance that is verified during development knockdown at the mRNA level as measured by qRT-PCR. The exact fold-repression is reported for each target and depends on gRNA design, promoter strength, and induction conditions.
Yes. Using dCas12a (Cpf1) allows processing of a single CRISPR RNA array to generate multiple gRNAs, enabling multiplexed repression. We routinely design arrays for 2–4 targets. For higher multiplexing, please discuss with our team during project scoping.
Our inducible CRISPRi system using β-estradiol provides reversible repression.Removal of the inducer leads to gradual recovery of target gene expression.
We perform colony PCR to verify construct integration or plasmid maintenance, Sanger sequencing of the gRNA cassette, and qRT-PCR (or fluorescence reporter assay) to quantify repression. Growth curves are measured to confirm no unintended fitness defects.
Yes. We can adapt the CRISPRi system to polyploid industrial strains, though transformation efficiency and repression levels may differ from haploid laboratory strains. Compatibility is assessed during the initial consultation.

Scientific Literature

References

Momen-Roknabadi A, Oikonomou P, Zegans M, et al. An inducible CRISPR interference library for genetic interrogation of Saccharomyces cerevisiae biology. Communications biology. 2020;3(1):723. View on PubMed
Ciurkot K, Gorochowski TE, Roubos JA, et al. Efficient multiplexed gene regulation in Saccharomyces cerevisiae using dCas12a. Nucleic acids research. 2021;49(13):7775-7790. View on PubMed
Feng Q, Li J. Construction and Validation of a CRISPR/dCpf1-Based Transcriptional Regulatory System in Saccharomyces Cerevisiae. Studies in health technology and informatics. 2023;308:680-688. View on PubMed
Spavieri JM, Inacio TG, Seguchi G, et al. Impact of CRISPRi-Mediated Titration of GPD Genes on the Fermentative Performance of S. cerevisiae. ACS synthetic biology. 2025;14(11):4412-4423. View on PubMed

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