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Objective: Produce correct oligo at correct length for application. Considerations: Length of oligo, Dimethoxytrityl (DMT) protection group on or off, synthesis system, purification system, and filtration system.

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• Solid-Phase Synthesis Objective: Cleavage from solid support and optional removal of protection groups.
• Considerations: Treatment in 25–28% ammonia at elevated temperature, additional cleavage step for RNA, and need for resistant bottles.
• Deprotection of nucleotide protection groups: Choose appropriate linker for the solid support, cleavage protocols for DNA or RNA, and develop process to treat column content with ammonia solution and elevated temperature.
• Obtaining full-length oligonucleotides: Different purification routes for DMT-ON or DMT-OFF.
• Strategies: DMT on and use hydrophobic interaction chromatography (HIC) or DMT off and use ion exchange chromatography (IEX).
• Separation of full-length oligonucleotides from modified oligonucleotides and short-mers with DMTr off.

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