I would like to synthesize the following oligo: 5′-CAGGGGCCTACACGTTGTA-3′. The problem I understand is the 4 Gs (starting at base 3) in a row will potentially cause synthesis issue. Also this G-quadruplex will potentially form undesirable secondary structure for PCR applications in the presence of potassium. To mitigate, I learned that it’s possible to synthesize an oligo with a G analog, such as 7-Deaza-2′-deoxyguanosine to lessen the chance for the potential problems mentioned above. If this analog indeed is the best course of action, what’s the best position(s) to insert this G analog, and how many to insert? However, I also read inserting multiple 7-Deaza-2′-deoxyguanosine could be problematic because 7-deaza-dG is sensitive to the iodine-based oxidizer solution used in phosphoramidite-based DNA synthesis. So, a suggestion is to substitute 7-deaza-dG with 7-deaza-8-aza-dG, which I’m not sure you have available as an option. -Chun-Nan

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Dear Chun-Nan,

Thank you for your interest in TriLink. Poly guanosine and deoxyguanosine stretches have the propensity to form tetraplexes (complexes consisting of four different strands all bound at the poly G region). These complexes are very tight and make oligonucleotide synthesis difficult.

The use of 7-deaza-dG during PCR reduces the secondary structure but, unfortunately, the 7-deaza-dG and 7-deaza-8-aza-dG amidites have the same innate properties as guanosine during oligonucleotide synthesis. Synthetic and processing capabilities would still be problematic. For multiple incorporations of 7-deaza-dG, CSO is recommended as the oxidizing solution to combat degradation.

TriLink has devised specialized protocols for multiple and/or consecutive insertions of guanosine or 7-deaza guanosine. Due to the complexity of manufacturing, there is typically reduced yield or purity compared to a sequence with better base distribution. The number of incorporations of guanosine, 7-deaza-guanosine and/or 7-deaza-8-aza- guanosine will determine the final effect.

I would recommend trying the unmodified compound before proceeding with the modified version as the cost of the modifications will lead to an overall higher price.

Best regards,
Katelyn Murphy
Product Manager