Oligonucleotide Synthesis: Advantages of Solid-phase Synthesis

Oligonucleotide Synthesis 

The chemical synthesis of relatively short segments of nucleic acids with a defined chemical structure is known as Oligonucleotide Synthesis (sequence). The method is particularly valuable in today's laboratory since it allows for quick and low-cost access to custom-made oligonucleotides of the appropriate sequence.

Every year, millions of oligonucleotides are produced for use in laboratories all around the world. Small amounts of DNA are required for most applications, and Oligonucleotide Synthesis is generally done at the 40 nmol scales or lower. Most biochemical and biological studies can be done with this amount of material. Much greater amounts of DNA (10 mol or more) can be synthesized for biophysical research (NMR and X-ray crystallography), and solid-phase technologies have been developed to allow the synthesis of multi-kilogram quantities of oligonucleotides for usage as therapeutic molecules in the extreme (e.g. antisense oligonucleotides). Oligonucleotides are synthesized for all of these reasons. Oligonucleotides are nearly entirely synthesized utilizing automated solid-phase processes for all of these functions.

Advantages of solid-phase synthesis

• Peptide synthesis, Oligonucleotide Synthesis, oligosaccharide synthesis, and combinatorial chemistry all involve solid-phase synthesis.
• Solid-phase synthesis takes place on a solid substrate held between filters in columns that allow all reagents and solvents to easily pass through. There are several advantages to solid-phase synthesis over solution synthesis.
• Large amounts of solution-phase reagents can be employed to speed up processes; contaminants and excess reagents are washed away, and no purification is necessary after each stage; the process can be automated using computer-controlled solid-phase synthesizers.

Controlled-pore glass (CPG)

The Oligonucleotide Synthesis takes place in controlled-pore glass, which is stiff and non-swelling with deep pores. Glass supports with 500 (50 nm) pores are mechanically strong and commonly employed in the synthesis of short oligonucleotides. When oligonucleotides longer than 40 bases are produced on resins with a pore size of 500, however, synthesis yields plummet substantially. Because the expanding oligonucleotide obstructs the pores, the chemicals' diffusion through the matrix is reduced. Despite the fact that large-pore resins are more fragile, 1000 CPG resin has proven to be enough for the synthesis of oligonucleotides up to 100 bases in length, while 2000 supports can be used for longer oligonucleotides.

Polystyrene (PS)

Highly cross-linked polystyrene beads have good moisture exclusion qualities and can be used to make oligonucleotides quickly, especially on a small scale.

Solid supports for traditional Oligonucleotide Synthesis are commonly made with a nucleoside loading of 20-30 mol per gram of resin. Due to steric hindrance between adjacent DNA chains attached to the resin, synthesis of oligonucleotide becomes less efficient at higher loadings; however, polystyrene supports with loadings of up to 350 mol / g are used in some applications, particularly for short oligonucleotides, and enable the synthesis of large quantities of oligonucleotides.