Custom Strain Engineering

Corynebacterium glutamicum Genome Editing & Metabolic Engineering Solutions

Accelerate your strain development with CRISPR-based genome editing and metabolic pathway redesign in C. glutamicum.

CRISPR-mediated gene knockouts, integrations, and point mutations
Metabolic pathway reconfiguration for amino acids, organic acids.
Transformant selection, clonal validation by PCR and Sanger.

End-to-End Capability

From gRNA design to shake-flask evaluation, our integrated workflow delivers validated edited clones with clear documentation for downstream development.

In silico gRNA design and off-target prediction
Transformation and colony screening
Sequencing-confirmed clonal isolates

CRISPR-Based Editing

Pathway Redesign

Fermentation Testing

Our Approach

Precision Genome Editing for C. glutamicum

We combine established CRISPR tools with metabolic engineering know-how to deliver tailored production strains. Every project begins with a design consultation, ensuring edits align with your target compound and process goals.

CRISPR-Mediated Modifications

Leverage Cas9 or Cas12a systems for targeted gene knockouts, precise integrations, and base edits. Custom guide RNAs and repair templates are designed for high editing efficiency.

GRNA design with in silico specificity scoring
Donor template construction (ssDNA or dsDNA)
Transformation and selection on marker plates

Metabolic Pathway Reconfiguration

Redesign native pathways to redirect flux toward desired products. We evaluate candidate gene targets based on host metabolism and literature, then implement edits iteratively.

Pathway flux analysis and bottleneck identification
Gene overexpression, downregulation, or deletion
Shake-flask production screening

Design & Simulation

In silico design of editing strategies including gRNA specificity, homologous arms, and integration loci. Off-target sites are predicted and minimized.

Editing & Selection

Transformation of C. glutamicum with CRISPR plasmids or ribonucleoproteins. Edited clones are selected via antibiotic markers or counterselection systems.

Validation & Handover

PCR screening, Sanger sequencing, and plasmid curing to confirm stable edits. A detailed report with sequence traces and growth data is provided.

What You Receive

Comprehensive Strain Handover

Each project delivers validated clonal isolates with full experimental documentation to support your next development stage.

Clone & Data Package

Purified, sequence-verified edited clones (as glycerol stocks)
Sequencing electropherograms confirming the intended edit(s)
PCR gel images and colony screening results
Plasmid maps and gRNA/template sequences (when applicable)

Supporting Documentation

Experimental design summary and workflow timeline
Off-target analysis report (in silico + optional targeted amplicon sequencing)
Growth and production data from shake-flask cultures (when scoped)
Quality control checklist and storage recommendations

Project Boundaries

Case-by-Case Customization

Every project is scoped individually based on target genes, desired edits, and production metrics. The table below lists typical parameters discussed during the design phase.

Parameter	Typical project scope
Host Strain	Common wild-type or derived strains (e.g., ATCC 13032, ATCC 13869); custom strains as agreed
Edit Type	Gene knockout, point mutation, short insertion, gene integration (size and locus scoped)
Number of Edits per Clone	Single or multiplex (up to 3 simultaneous edits typical; more evaluated case by case)
gRNA Design	In silico design with specificity analysis; up to 5 gRNAs per target tested, top performer selected
Transformation Method	Electroporation of competent cells; optimization of conditions for high efficiency
Clonal Validation Depth	PCR screening (colony PCR) + Sanger sequencing of target region; optional whole plasmid sequencing
Metabolic Analysis	Shake-flask fermentation with basic metabolite quantification (HPLC or enzymatic assays); additional analyses scoped per project
Fermentation Scale (Testing)	Scoped per project after consultation
Turnaround Time	Scoped per project after consultation
Technical Support	Named scientific contact at project start; milestone review calls; email response within 1 business day

Process Steps

Standard Project Workflow

Our five-stage engagement ensures transparent communication and rigorous quality control at every step.

Consultation & Design

We review your target genes, desired edits, and production goals. A feasibility assessment is performed and a detailed experimental design document is prepared for your approval.

Plasmid & Donor Construction

Custom CRISPR plasmids (encoding Cas9 or Cas12a) and repair templates are assembled. Constructs are sequence-verified before proceeding to transformation.

Transformation & Selection

C. glutamicum cells are transformed via electroporation with the editing plasmids. Transformants are selected on antibiotic-containing plates and isolated for screening.

Clonal Validation

Colony PCR is used to identify positive clones. The edited region is Sanger sequenced to confirm the intended modification. Correct clones are cured of editing plasmids.

Strain Handover & Report

Validated clonal isolates are provided as glycerol stocks. A project report including methods, QC, and datasets including sequencing data, growth characteristics, and quality control results is delivered.

Use Cases

Where Our Service Adds Value

C. glutamicum is a proven industrial workhorse. Our genome editing service supports a range of metabolic engineering objectives.

Amino Acid Overproduction

Engineer strains for high-yield production of L-lysine, L-threonine, L-tryptophan, and other amino acids by relieving feedback inhibition and redirecting metabolic flux.

Knockout of competing pathway genes
Introduction of feedback-resistant enzyme variants
Shake-flask validation of titer and yield

Organic Acid & Diol Production

Evaluate production of succinic acid, lactic acid, and 1,3-propanediol through targeted gene deletions and overexpression of rate-limiting enzymes.

Deletion of byproduct-forming enzymes
Integration of heterologous pathways
Growth and production profiling under controlled conditions

Bioproduct & Chemical Synthesis

Expand the product portfolio to include polyamides, carotenoids, and other high-value chemicals by introducing non-native biosynthetic modules.

Design of synthetic operons for new pathways
Genomic integration of multiple genes
Preliminary product quantification and pathway debugging

Common Questions

Frequently Asked Questions

Addressing typical concerns about timeline, deliverables, and technical feasibility.

Editing efficiency varies by target locus, editing type, and strain background. Knockout efficiencies typically range from project-dependent performance that is verified during development to project-dependent performance that is verified during development in our hands. For knock-ins and point mutations, efficiency is lower and project-dependent. During the design phase we provide an estimated success rate based on similar projects.
We perform PCR screening and Sanger sequencing of the target region after curing of editing plasmids to confirm stability. For off-target analysis, we use in silico prediction tools and, optionally, targeted amplicon sequencing of top predicted off-target sites. Whole-genome sequencing is not included by default but can be scoped.
Turnaround is a timeline confirmed during project scoping, depending on the complexity of the edits and the number of validation steps required. Multiplex edits or projects requiring additional metabolic analysis may extend the timeline. A detailed schedule is provided during project initiation.
Yes. We can accept proprietary strains under a confidentiality agreement. The strain must be non-pathogenic and amenable to electroporation. We will need a small sample for competence testing. All work is performed using the provided strain and no third-party distribution occurs.

Scientific Literature

References

Wang Y, Zheng P, Sun J. [Recent advances in developing enabling technologies for Corynebacterium glutamicum metabolic engineering]. Sheng wu gong cheng xue bao = Chinese journal of biotechnology. 2021;37(5):1603-1618. View on PubMed
Liu Y, Liu Y, Zheng P, et al. Cytosine Base Editing in Bacteria. Methods in molecular biology (Clifton, N.J.). 2023;2606:219-231. View on PubMed
Park E, Kim HJ, Seo SY, et al. Shikimate Metabolic Pathway Engineering in Corynebacterium glutamicum. Journal of microbiology and biotechnology. 2021;31(9):1305-1310. View on PubMed
Liu J, Liu M, Shi T, et al. CRISPR-assisted rational flux-tuning and arrayed CRISPRi screening of an L-proline exporter for L-proline hyperproduction. Nature communications. 2022;13(1):891. View on PubMed
Li C, Swofford CA, Rückert C, et al. Optimizing recombineering in Corynebacterium glutamicum. Biotechnology and bioengineering. 2021;118(6):2255-2264. View on PubMed

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