Glen Research provides efficient and inexpensive purification cartridges for DNA and RNA oligonucleotides that can purify up to 60mer and 150mer oligonucleotides using the RNA and DNA cartridges, respectively. The purification mechanism for these cartridges relies on the DMT-ON purification concept, where the desired full-length oligonucleotide with the hydrophobic DMT group sticks to the cartridge’s resin while the shorter, less hydrophobic failures elute out of the cartridges during the initial purification steps.1,2
The standard format of the DNA and RNA cartridges is the 1 umol scale version (Figure 1). Physically, they share the same dimensions (65 x 10mm) and can be used as single columns or in a 96-well plate format. What makes them different is the resin type and the amount of resin present, 150mg in the DNA cartridge versus 100mg in the RNA cartridge.
In recent years, we have heard anecdotally from customers that the DNA Glen Pak is also suitable for RNA. The DNA and RNA columns were developed separately, and we had not considered using the DNA version for RNA purification. As such, we decided to give it a try.
We tested the purification of RNA oligonucleotides of a couple of different lengths using our standard Glen-Pak RNA cartridge (60-6100) and its DNA equivalent (60-5100). The RNA oligonucleotides were synthesized in-house, and all the steps prior to the purification step (RNA synthesis, base deprotection, and 2’ deprotection) followed our TBDMS-Protected RNA guide and Glen-Pak cartridge purification guide (p26). RNA oligonucleotides were divided into two equal portions and loaded separately into the DNA and RNA cartridges. The RNA protocol (p26) was applied to both cartridges.2
The results were quite impressive. For a crude 21mer RNA oligonucleotide of 76.7% purity, the Glen-Pak DNA cartridge produced 97.5% purity compared to 98.5% for the Glen-Pak RNA cartridge. The obtained yields were also comparable at 63.5% and 66.5%, respectively. We also repeated these tests for 56mer RNA oligonucleotides and obtained similar results. Based on this data, we believe the Glen-Pak DNA cartridge is effective for any RNA sequence up to at least 56mer.
Figure 2. HPLC chromatograms for crude and cartridge-purified RNA. Data correspond to 21mer RNA oligonucleotide (5’- GGCUCCCCUCAACAACUUCTT-3’) before and after purification with either a Glen-Pak DNA or RNA cartridge.
The data obtained from this evaluation are very encouraging, and researchers can potentially stock and maintain just the DNA cartridges for both RNA and DNA oligonucleotide purification needs. Also, it should be noted that although we only tested the 150mg cartridge, we believe the other DNA Glen-Pak formats should work fine with RNA. Customers should be able to purify larger and smaller scales of syntheses on the 3g and 50mg cartridges, respectively, by scaling the RNA purification protocol accordingly.
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