Oligonucleotides Purification possibilities - BioMolecules

Purification possibilities

The chemical synthesis of oligonucleotides, according to the phosphoramidite method, is never 100% effi cient. All producers of phosphoramidites guarantee an average coupling efficiency of ~99% when coupling two nucleotides. This means that during the synthesis, the crude mixture also contains truncated failure sequences. In general, the required purity of the oligonucleotide is dependent on its application. Sometimes it is essential that only full length oligonucleotides (n) are present and sometimes the presence of short oligonucleotides (n-1, n-2, …) has no effect on the experimental results. Oligonucleotides with a length to 50 bases are purifi ed by us without additional costs using Solid Phase Extraction. An optional HPLC purifi cation, desalting, or PAGE2 gel purifi cation is also possible.

The Solid Phase Extraction (SPE) technology: We have developed a full automatic purification technology (SPE) for standard oligonucleotides (not modified, 15 – 50 bases long). This automated process allows the purification under salt-free conditions and is comparable with reversed phase cartridge purification. The SPE deletes every unusable product of the chemical synthesis, e.g. capping products. The result is an oligonucleotide free of impurities. The SPE method is automated which makes it possible to produce large quantities of oligonucleotides with a purity comparable to HPLC oligonucleotides. This is quicker, because the oligonucleotides are purified during the production process itself and additional HPLC purification is not required.

HPLC purification: For some applications HPLC purification is required. Depending on the type of oligonucleotide, purification is performed with reversed phase or anion-exchange. The capacity and resolution of HPLC columns are higher than SPE columns. That is why we advice a HPLC purification for large synthesis scales (>1 micromole), modified oligonucleotides and oligonucleotides longer than 50 bases.

PAGE2 purification: Gene synthesis with longer oligonucleotides is anticipated to become the industry gold standard due to the cost and efficiency improvements brought by the availability of building blocks of high purity and improved manufacturing processes. In addition to HPLC, we now offer high level purification of oligonucleotides using a very efficient purification technology developed by Elchrom Scientific AG.


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