Oligonucleotide Separation Technology: Synthesis Challenges and HPLC Isolation Options

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Martin Gilar, Bill Warren
Waters Applications Note
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Application Notes
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Application Notes
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The synthesis of oligonucleotides is a highly efficient process. However, small amounts of impurities are created in each cycle of synthesis. Understandably, longer oligonucleotides are more contaminated than short ones. The failure products, typically labeled N-1, N-2..., N-x, are prematurely halted shorter oligonucleotides. Some are missing a nucleotide(s) in the middle of sequence, rather than at the end. These products are called mismatch failure sequences. Some by-products of synthesis may have greater molecular weight (often labeled N+x) than the target oligonucleotide. This is a result of incomplete post-synthesis deprotection, or due to the branching of an oligo backbone during the synthesis. For labeled oligonucleotides, the failure products are also generated by failure to conjugate the label with the target sequence. This application note addresses how Waters® Oligonucleotide Separation Technology Columns is the most viable option for handling the challenges of purification and isolation of synthetic oligonucleotides.


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