Advantages of Liquid-phase Chemical Synthesis Method

The liquid-phase technique provides an alternative to the extensively employed solid-phase oligonucleotide synthesis method. In this strategy, a soluble polymeric carrier integrated into the reaction medium acts as the scaffold for elongating the chain. Afterward, the polymer is separated from unused reagents and soluble by-products using filtration or crystallization techniques. This approach eliminates any inconsistency resulting from support insolubility, thus enhancing the potential for process upscaling. This is particularly pertinent now as synthetic oligonucleotides are transitioning from research settings to practical biomedical applications. Consequently, the scientific community is actively exploring the development and evaluation of a more user-friendly liquid-phase chemical synthesis method for oligonucleotides. An innovative strategy known as high-performance liquid phase (HELP), utilizing soluble polyethylene glycol (PEG) as a polymeric support and employing phosphotriester chemistry for the coupling reaction, has demonstrated promising outcomes.

Key Technologies

Our liquid-phase synthesis platform is built upon a modified phosphoramidite chemistry, the most widely used technique for DNA and RNA synthesis. Unlike solid-phase synthesis, which uses an insoluble support, our liquid-phase oligonucleotide synthesis method utilizes a soluble polymeric support that remains dissolved in the reaction solvent. This homogeneous reaction environment provides several key advantages, including better reagent accessibility and more predictable, linear reaction kinetics. This approach allows for a reduction in the large excesses of expensive amidite monomers typically required in solid-phase methods to drive reactions to completion. Our innovative "one-pot" liquid-phase oligonucleotide technique for DNA synthesis allows for sequential coupling, oxidation, and deprotection in a single reaction vessel, followed by a single precipitation step, drastically streamlining the process and reducing overall solvent consumption.

Liquid-Phase Precision, Nucleotide by Nucleotide

CD BioGlyco commits to providing global clients with industry-leading Custom Carbohydrate Synthesis services, such as Custom Sugar-nucleotides Synthesis services. The HELP-based Synthesis of Oligonucleotides we provide mainly includes 3 key steps: functionalization of PEG, oligonucleotide assembly, deprotection & purification.

• PEG Functionalization
  The initial nucleoside is attached through PEG, which is linked to a symmetrical anhydride derivative of four 5'-O-dimethoxytrityl (DMT)-2'-deoxynucleoside-3'-O-succinates. The PEG-nucleosides are then subjected to analysis using ¹H-NMR spectroscopy. To confirm the complete removal of reagents, TLC is initially employed, followed by product characterization. Any residual unreacted PEG molecules are treated with acetic anhydride.
• Oligonucleotide assembly
  The process of oligonucleotide assembly is executed through detritylation, condensation, capping, and oxidation steps, adhering to established procedures.
• Deprotection & purification
  Upon the synthesis's completion, the oligomers are separated from the polymer using standard deprotection techniques. Most of the product is detached from PEG through oximation, succeeded by comprehensive hydrolysis using concentrated ammonia. Partially deprotected oligonucleotides are subjected to analysis via HPLC. Finally, after full deprotection with acetic acid, the product undergoes purification through ion exchange chromatography.

Workflow

Fig.1 Process of HELP for oligonucleotide synthesis. (CD BioGlyco)

Publication Data

Applications of Liquid-phase Chemical Synthesis Method

• Biomedical research: The liquid-phase synthesis method is extensively used to produce custom oligonucleotides for various research purposes, such as investigating gene expression, studying genetic variations, and exploring nucleic acid-protein interactions.
• Diagnostic assays: Oligonucleotides synthesized using the liquid-phase method are crucial components of diagnostic assays, including polymerase chain reactions (PCR), reverse transcription PCR (RT-PCR), and nucleic acid hybridization techniques.
• Next-generation sequencing (NGS): Oligonucleotides produced through the liquid-phase approach serve as primers and adapters in NGS technologies. They facilitate the high-throughput sequencing of DNA and RNA, enabling comprehensive genomic and transcriptomic analyses.

Advantages

• Liquid-phase oligonucleotide synthesis employs a uniform reaction mixture, enabling potentially boundless scalability.
• Oligonucleotides are synthesized using the liquid-phase technique and hold the potential for advancing extensive drug development efforts.
• The liquid-phase approach to oligonucleotide synthesis is expected to contribute to the progression of large-scale fabrication of DNA materials.

Frequently Asked Questions

Associated Services

Liquid-phase chemical synthesis for oligonucleotides, leveraging soluble polymer supports (e.g., PEG) and homogeneous reaction conditions, enables scalable, cost-effective production of DNA/RNA sequences with reduced reagent consumption and streamlined purification via precipitation or membrane filtration. Forwarding glycotherapeutic vectors and nucleic-acid-conjugate integrations, this platform necessitates co-delivered-associated enzymatic services: concerted enzymatic catalysis fabricates nucleotide sugar donors satisfying exacting purity specifications.

At CD BioGlyco, we have established the high-quality Glyco™ Synthesis Platform that is dedicated to glycobiology research. Our commitment to excellence is evident in every aspect of the Chemical-based Oligonucleotide Synthesis Service. Our team is dedicated to maintaining the highest standards and ensuring client satisfaction, particularly in the realm of liquid-phase chemical synthesis methods for oligonucleotides. If you are interested in our services, please do not hesitate to contact us for more details.