Oligo Pool Synthesis Technologies: In Situ vs. Array-Based Manufacturing

The rapid advancement of oligo pool synthesis technologies has become a cornerstone of synthetic biology, enabling applications such as gene library construction, CRISPR screening, and precision genome editing. As a leader in this field, Dynegene Technologies has pioneered high-throughput DNA synthesis solutions since its establishment in 2018, leveraging proprietary platforms like its in situ synthesis technology to deliver industry-leading accuracy and scalability. This article compares two dominant methodologies—in situ synthesis and array-based manufacturing—highlighting their technical distinctions, applications, and alignment with Dynegene’s innovations.

Core Technical Principles

In Situ Oligo Synthesis

In situ synthesis builds oligonucleotides directly on solid substrates (e.g., glass or silicon) using spatially controlled chemical reactions. Dynegene’s proprietary high-throughput synthesis platform exemplifies this approach, achieving ultra-high-density synthesis with parallelized light-activated deprotection. Key advantages include:

1.     Error Minimization: Real-time monitoring and closed-loop quality control ensure high fidelity, critical for CRISPR guide RNA libraries and custom modifications.

2.   Flexibility: Supports site-specific additions such as biotin tags or fluorescent markers for downstream applications like pull-down assays.

Applications:

• CRISPR Screening: Synthesis of >100,000 unique sgRNAs for genome-wide studies.
• Diagnostic Probes: Custom oligo pools for molecular diagnostics and companion diagnostics.

Array-Based Oligo Synthesis

Array-based methods pre-synthesize oligos on microchips followed by enzymatic amplification. This approach is ideal for large-scale projects due to:

1.     Scalability: Production of millions of oligos per batch, suitable for NGS panels or synthetic gene libraries.

2.   Cost Efficiency: Bulk purification reduces per-oligo costs for projects exceeding 10,000 sequences.

Advantages:

• Uniformity: Batch consistency ensures minimal sequence bias, vital for mRNA vaccine template synthesis.
• Length Optimization: Supports oligos up to 300 nt, sufficient for most HDR template designs.

Technical Challenges and Solutions

Error Rates and Validation

• In Situ: Error rates (~1/500 nt) are mitigated via enzymatic mismatch repair (e.g., T7 Endonuclease I) and NGS validation.
• Array-Based: Rigorous batch QC ensures <0.3% error rates, meeting standards for therapeutic-grade oligos.

Length Limitations

• In Situ: Achieves 200–230 nt lengths, ideal for sgRNA and diagnostic probes.
• Array-Based: Optimized workflows extend synthesis to 300 nt for gene assembly.

Emerging Trends (2025)

1.     Automated Synthesis: Machine learning-driven error correction for yield optimization.

2.   Cell-Free Systems: Integration with TX-TL platforms for rapid metabolic pathway prototyping.

Conclusion

Dynegene’s in situ synthesis and array-based platforms cater to diverse needs:

• In Situ: Custom libraries with tailored modifications for diagnostics and CRISPR.
• Array-Based: High-throughput projects requiring cost-effective, uniform synthesis.

With a 10,000 m² production facility and partnerships with over 1,000 clients globally, Dynegene continues to drive innovation in synthetic biology. Explore our Oligo Synthesis Solutions or contact our team for project-specific guidance.