Oligonucleotide synthesis plays a crucial role in molecular biology, underpinning advancements in areas such as polymerase chain reactions (PCR), gene editing, DNA sequencing, and therapeutic delivery. Over the past 35 years, this process has driven significant progress in diagnostics, biotechnology, and pharmacology, thanks to advancements in oligonucleotide synthesis technology.
In this article, we delve into the oligonucleotide synthesis process and its impact on modern medicine.
What is an Oligonucleotide?
Oligonucleotides are short segments of nucleic acids, either single or double-stranded, that form single-chain biological polymers. Each nucleotide in an oligonucleotide consists of a nitrogenous base, a five-carbon sugar (forming a nucleoside), and one or more phosphate groups. These nucleotides can also include non-natural or modified bases, such as locked nucleic acids (LNA) and Morpholino, or structural modifications to their backbones.
These nucleotide sequences are essential for creating RNA and DNA molecules, with RNA being a single-stranded biopolymer containing ribose sugar and DNA a double-stranded molecule with deoxyribose sugar. The sequences of these nucleic acids are fundamental to numerous biological, medical, forensic, and clinical applications.
What is Solid-Phase Oligonucleotide Synthesis and How Does it Work?
In modern medicine, the phosphoramidite chemistry method, developed over 35 years ago, is widely used for oligonucleotide synthesis. This method, known for its simplicity and efficiency, underpins automated oligonucleotide synthesis systems.
The process starts with anchoring the first nucleotide to a solid support, typically a resin. Subsequent nucleotides are added sequentially, ensuring precise control over the sequence. The solid support facilitates purification and automation of the process.
The synthesis involves a repetitive cycle of four steps: deblocking, coupling, capping, and oxidation. These steps extend the oligonucleotide chain in a controlled manner. After achieving the desired sequence, the oligonucleotide is cleaved from the solid support and undergoes post-synthesis processing, including deprotection.
What are the Alternative Methods of Oligonucleotide Synthesis?
Solid-phase synthesis can face challenges, such as decreased yield with increasing chain length. Alternative methods include:
- Solution-Phase Synthesis: Nucleotides are combined in solution without solid support. This method supports larger-scale reactions but poses challenges in purification and yield.
- Enzymatic Synthesis: Uses enzymes like DNA polymerases to synthesize oligonucleotides. It is highly selective and capable of producing longer sequences but requires specific enzymes and is prone to errors.
- Array-Based Synthesis: Simultaneously synthesizes multiple oligonucleotides on a solid surface, such as a microarray. This method is advantageous for high-throughput applications but may have limitations in sequence length and yield.
- Phosphite-Triester Method: Uses solution-phase chemistry with nucleotide phosphoramidites and phosphites to form oligonucleotides. Although less common, it offers an alternative to solid-phase synthesis.
Each method has its benefits and limitations, and the choice depends on factors like scalability, sequence length, and specific application requirements.
Why Choose Bachem?
At Bachem, we offer expertise in oligonucleotide process development, GMP production, and regulatory compliance. Our long-standing commitment to innovation and our extensive experience enhance every phase of the process, maximizing your product’s potential. We utilize advanced engineering solutions for precise process control and high throughput, ensuring your product reaches the market effectively.
Our team specializes in developing and validating test methods for large and complex molecules. We excel in solid-phase synthesis, protecting group chemistry, chromatography-based purification, and advanced techniques like ultrafiltration, diafiltration, precipitation, and lyophilization.
Leverage our cutting-edge chemistry, manufacturing, and control (CMC) services for oligonucleotide-based APIs. We support scalable manufacturing processes and assist with IND and NDA applications.
Elevate your projects with Bachem’s unparalleled oligonucleotide manufacturing expertise. Contact us to learn more about our innovative solutions, precision in synthesis, and commitment to quality.
