In the rapidly evolving field of synthetic glycobiology, the development of robust and high-performing microbial or mammalian chassis is the cornerstone of successful bioproduction. However, the genetic complexity of these engineered hosts, often subjected to extensive gene editing, metabolic rewiring, and the insertion of heterologous glycosylation pathways, demands a validation method of unparalleled resolution. Whole genome sequencing (WGS) has emerged as the definitive tool for this purpose. Unlike traditional polymerase chain reaction (PCR)-based or targeted sequencing methods that only look at specific loci, WGS provides a single-nucleotide resolution map of the entire genome.
At CD BioGlyco, we recognize that even a minor off-target mutation or an unintended genomic rearrangement compromises yield and product quality. Our WGS-based chassis validation service is designed to provide scientists with a high-fidelity "genetic audit" of their engineered strains. By leveraging state-of-the-art sequencing platforms and specialized bioinformatic pipelines, we ensure that your chassis is exactly what you designed it to be, providing the confidence needed to move from the laboratory to large-scale fermentation.
Key Technologies
Next-Generation Sequencing (NGS)
We utilize the Illumina NovaSeq and MiSeq systems to generate high-coverage, short-read data. This technology is the gold standard for identifying single-nucleotide polymorphisms (SNPs) and small insertions/deletions (InDels) with extreme precision. It is essential for verifying that specific codon-optimized glyco-enzymes have been accurately integrated into the host genome.
Third-Generation Sequencing (TGS)
Long-read sequencing is critical for chassis validation, particularly when dealing with repetitive regions or large-scale structural variants (SVs). Our high fidelity allows us to resolve complex genomic rearrangements, large inversions, and translocations that short-read platforms often miss. This ensures the structural integrity of the entire chassis architecture.
Integrated Bioinformatics Pipeline
Raw sequencing data is processed through our proprietary bioinformatic suite. This includes de novo assembly for novel chassis strains and reference-based mapping for well-characterized hosts like Escherichia coli or Saccharomyces cerevisiae. Our algorithms are optimized for variant calling, functional annotation, and comparative genomics to highlight any deviations from the "design" sequence.
Our scope focuses on the absolute verification of the genetic makeup of engineered hosts used in glycobiology research and industrial production. Whether you are working with bacterial, fungal, or mammalian cell lines (such as CHO cells), we provide an analysis that spans from the primary sequence to higher-order genomic structures.
This service is particularly vital for validating the success of complex "glyco-engineering" projects where multiple genes are deleted or inserted to create human-like glycosylation patterns. We perform high-depth sequencing to detect subclonal mutations that might arise during the subculturing of engineered strains. We provide a comparative analysis between the parent strain and the engineered derivative to confirm that no unintended "genomic scars" remain from the editing process. Our validation includes the verification of selection markers, promoter stability, and the presence of any endogenous mobile genetic elements that could destabilize the production strain over time.
By integrating WGS into your development cycle, CD BioGlyco helps you mitigate the risks of phenotypic instability and ensures that your biological chassis is a stable, reliable platform for the synthesis of high-value glycoconjugates.
Workflow
Genomic DNA Extraction
The process begins with the cultivation of the client's chassis strain under optimized conditions. We then perform high-molecular-weight (HMW) genomic DNA (gDNA) extraction. This preserves the integrity of the DNA fragments, which is paramount to ensure the assembly of a "closed" circular genome.
Library Preparation
Following extraction, the gDNA undergoes rigorous quality control (QC) using fluorometric quantification (Qubit) and fragment size analysis. We offer multiple library types, including PCR-free libraries for Illumina sequencing and SMRTbell libraries for PacBio.
High-Throughput Sequencing
Samples are loaded onto the appropriate platform based on the project requirements. We typically recommend a minimum of 50x to 100x coverage for microbial chassis to ensure high-confidence variant calling. For complex polyploid fungal or mammalian hosts, higher coverage depths are employed.
Genome Assembly and De Novo Mapping
Our bioinformaticians perform de novo assembly to reconstruct the genome from scratch or utilize reference-guided mapping. This step allows us to identify the exact location of all genetic modifications and verify that the synthetic constructs are correctly integrated into the designated loci.
Variant Calling and Structural Analysis
We employ advanced algorithms to detect SNPs, InDels, and SVs. This is the core of the validation service, where we compare the sequenced genome against the user's design. We specifically look for off-target edits and chromosomal instabilities that could affect the fitness of the chassis.
Delivery Reporting
The final deliverable is an exhaustive validation report containing assembly statistics, variant lists, and functional annotations. More importantly, our specialists provide a consultation to interpret the findings, helping you decide whether the strain is ready for scale-up or requires further optimization.
Publication Data
DoI: 10.3390/microorganisms14010154
Journal: Microorganisms
IF: 4.2
Published: 2026
Results: This study presents a genomic and pathogenic characterization of Pasteurella multocida strain P6, a capsular serotype A isolate from a Tan sheep that died of respiratory disease in Ningxia, China. WGS reveals a 2,289,251 bp genome (40.2% GC content) encoding 2155 genes, plus mobile genetic elements (genomic islands, prophages, transposons, a gene editing locus) enhancing genomic plasticity. Phylogenetic analysis groups P6 closely with strains 166CV and 103220, distinct from rodent/avian isolates. A murine model confirms its virulence, causing severe pulmonary lesions. Functional analyses identify genes linked to carbohydrate metabolism, virulence (capsule/LPS biosynthesis, iron acquisition), and antimicrobial resistance. Comparative genomics with reference strain PM70 shows conserved core functions but enriched mobilome-associated genes in P6. This case-based study provides baseline genomic data for P. multocida in Tan sheep, highlighting its adaptive and pathogenic traits, and supports future disease surveillance and control efforts.
Fig.1 Circular genome map of Pasteurella multocida P6. (Zhao, et al., 2026)
Applications
Therapeutic Glycoprotein Production
Validating mammalian or microbial chassis for the production of monoclonal antibodies and erythropoietin, ensuring that the glycosylation-related genes are stable and correctly expressed.
Metabolic Engineering for Rare Sugars
Ensuring that multigene pathways for the synthesis of human milk oligosaccharides (HMOs) or other rare glycans are correctly integrated and free of deleterious mutations.
Vaccine Development
Verifying the genetic stability of attenuated or recombinant bacterial strains used as platforms for conjugate vaccines or O-antigen modification.
Industrial Enzyme Synthesis
Confirming the genomic integrity of high-yield fungal chassis (e.g., Pichia pastoris) used for the secretion of glycosidases and glycosyltransferases for industrial biocatalysis.
Advantages
Unrivaled Resolution
Our WGS service provides a 360-degree view of the genome, detecting single-nucleotide changes and large structural rearrangements that traditional QC methods inevitably miss.
Specialized Glyco-Focus
Unlike general sequencing providers, CD BioGlyco understands the specific genetic requirements of glycosylation pathways, allowing for more relevant functional annotations.
Hybrid Sequencing Technology
By combining short-read accuracy with long-read structural insights, we provide the most complete and "closed" genome assemblies available in the industry.
Tailored Bioinformatic Pipelines
We don't use a "one size fits all" approach. Our pipelines are customized to the specific biology of your host organism, ensuring higher sensitivity in variant detection.
Frequently Asked Questions
Customer Review
"The depth of analysis provided by CD BioGlyco's WGS service was instrumental in our recent project. We discovered a large-scale duplication in our yeast chassis that our standard PCR assays had missed for months. Their bioinformatic support is second to none."
– B.M., Senior Scientist
"Working with CD BioGlyco saved us significant time. The turnaround was faster than expected, and specialists provide a consultation to interpret the findings."
– R.U., Director of R&D
"They understood exactly why we were looking at specific glycosyltransferase loci and provided insights we hadn't even considered."
At CD BioGlyco, our WGS-based chassis validation service is more than just a sequencing run; it is a solution for genetic quality control in the age of synthetic biology. By providing the clarity and precision needed to verify engineered hosts, we empower our clients to push the boundaries of what is possible in glycobiology. Please feel free to contact us to help you design the perfect validation strategy.
Reference
Zhao, Y.; et al. Whole-genome sequencing and pathogenic characterization of a Pasteurella multocida serotype A isolate from a case of respiratory disease in Tan sheep. Microorganisms. 2026, 14(1): 154. (Open Access)
For research use only. Not intended for any clinical use.
Fig.1 Circular genome map of Pasteurella multocida P6. (Zhao, et al., 2026)
