Yeast Gene Editing

Yeast Gene Knock-in Service

Precise, customizable integration of genes or tags into the Saccharomyces cerevisiae genome using CRISPR/Cas9.

Precise genomic integration at user-defined.
Customizable donor design with homology arms
Marker-free and scarless integration options

Targeted Knock-in with Flexibility

Our service enables efficient insertion of genes, tags, or regulatory elements in S. cerevisiae, leveraging CRISPR/Cas9-mediated homology-directed repair for stable genomic integration. Using custom-designed guide RNAs and donor templates with homology arms, we deliver verified knock-in clones for your chosen locus. Marker-free integration options and flexible insert sizes are scoped to project requirements.

CRISPR/Cas9-mediated homology-directed repair
Scarless integration possible with marker-free donors
Applicable to a range of S. cerevisiae strains as scoped per project

Custom sgRNA Design

Homology Arm Donor Construction

Verified Clone Delivery

Approach

CRISPR/Cas9-Mediated Gene Knock-in

Combining guide RNA design, donor template construction, and transformation for precise integration.

Guide RNA and Donor Design

We design target-specific single guide RNAs and construct donor templates with homology arms flanking your gene of interest.

In silico sgRNA design for on-target efficiency
Donor template with customizable homology arms (typical length 300–500 bp each)
Optional selection markers or marker-free cassettes

Transformation and Clone Screening

Co-delivery of Cas9-sgRNA and donor into yeast, followed by selective plating and molecular screening for correct integration.

Chemical or electroporation transformation of S. cerevisiae
Colony PCR screening to identify integrants
Sanger sequencing verification of junction regions

Scarless Integration

Marker-free donor designs allow seamless knock-in without residual selection markers, suitable for metabolic engineering and synthetic biology.

Flexible Insert Size

Inserts ranging from short tags to multi-gene constructs can be integrated; larger fragments are scoped case by case.

Marker-Free Options

Recyclable or excisable markers enable repeated rounds of editing in the same strain without genomic footprint.

Deliverables

What You Receive

Comprehensive package of validated materials and data.

Physical Deliverables

Purified donor plasmid (if applicable)
Verified yeast clone(s) – glycerol stock or agar plate
Genomic DNA from confirmed knock-in clone
Detailed QC report (PCR gel images, sequencing chromatograms)

Data Deliverables

SgRNA and donor design files (sequence maps)
PCR confirmation results and primer sequences
Sanger sequencing data covering integration junctions
Integration site analysis summary

Scope

Customization Options

Parameter | Typical Project Scope

Parameter	Typical project scope
Target Locus	User-defined genomic site; evaluated for accessibility and homology arm design.
Insert Size	Up to several kb; larger inserts require case-by-case assessment.
Homology Arm Length	Scoped per project after consultation
Selection Marker	Scoped per project after consultation.
Number of Clones	Confirmed clones (quantity scoped per project).
Strain Background	S. cerevisiae laboratory or industrial strains upon data review.

Workflow

How It Works

A streamlined five-step process from design to delivery.

Project Consultation and Target Design

We discuss your target gene, integration site, and insertion requirements to define the optimal knock-in strategy and scope.

SgRNA and Donor Template Construction

Custom single guide RNA and donor DNA with homology arms are designed, synthesized, and cloned into appropriate vectors.

Yeast Transformation

The Cas9-sgRNA construct and donor template are co-transformed into S. cerevisiae using standard chemical or electroporation methods.

Clone Screening and PCR Verification

Transformants are plated on selective media; candidate colonies are screened by colony PCR to identify clones with correct integration.

Sequence Confirmation and Final Reporting

Positive clones are confirmed by Sanger sequencing across both integration junctions; a final report and materials are delivered.

Applications

Pathway Optimization

Introduce heterologous or modified genes into the yeast genome to support metabolic engineering objectives.

Introduce heterologous enzymes
Modify endogenous pathway genes
Integrate multiple genes in one locus

FAQ

Frequently Asked Questions

Common inquiries about our yeast gene knock-in service.

Knock-in efficiency varies with target locus, insert size, and homology arm design. Our service screens sufficient clones to deliver at least one confirmed integrant; exact rates are scoped per project.
Yes, we offer marker-free donor designs that allow integration without leaving a selection marker. This is achieved via co-transformation with a helper plasmid or counter-selection methods, depending on project requirements.
We perform colony PCR across both 5' and 3' junctions to confirm integration, followed by Sanger sequencing of the entire insert and flanking regions. A comparative analysis of the edited sequence to the intended design is provided.
Turnaround typically ranges from 6 to 8 weeks from project kickoff, depending on target complexity, insert size, and cloning steps. A detailed timeline is provided during the project scoping phase.

Scientific Literature

References

Li ST, Kamada K, Honda A, et al. LLP1 is a pyrophosphatase involved in homeostasis/quality control of dolichol-linked oligosaccharide. The Journal of cell biology. 2025;224(11). View on PubMed
Feng Y-H, Song J-Z, Zhu J, et al. Template plasmids optimized for deletion of multiple genes in yeast Saccharomyces cerevisiae. Microbiology spectrum. 2024;12(11):e0132024. View on PubMed
Li H, Ma W, Lyv Y, et al. Glycosylation Modification Enhances (2S)-Naringenin Production in Saccharomyces cerevisiae. ACS synthetic biology. 2022;11(7):2339-2347. View on PubMed
Jiang Y, Liang C, Zhao W, et al. Cell Cycle Progression Influences Biofilm Formation in Saccharomyces cerevisiae 1308. Microbiology spectrum. 2022;10(3):e0276521. View on PubMed

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