Despite the fact that Oligonucleotide Synthesis were once done in solution, the solid-phase approach quickly became the preferred method, especially for large-scale (5 to 15 mmol) preparations. To produce practical yields of full-length oligonucleotides, coupling yields per step must be as close to 100% as possible. For example, in the synthesis of a 10-mer oligonucleotide, the overall yields of the full-length product will be 65 percent, 78 percent, and 95%, resembling the step-wise coupling values of 90%, 97%, and 99%.
The phosphoramidite, H-phosphonate, and phosphotriester methods of solid-phase Oligonucleotide Synthesis became the methods of choice due to their high step-wise coupling yields and quick kinetics (1–2 min). The discovery of suitable solid substrates, as well as extremely efficient coupling reaction procedures, has facilitated the synthesis of oligonucleotides utilising automated DNA synthesisers in this context.
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