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High-throughput chemical modification of oligonucleotides for systematic structure-activity relationship evaluation.

High-throughput chemical modification of oligonucleotides for systematic structure-activity relationship evaluation.

Bioconjugate chemistry (2014-11-15)

Daniel Zewge, Francis Gosselin, Denise M Kenski, Jenny Li, Vasant Jadhav, Yu Yuan, Sandhya S Nerurkar, David M Tellers, W Michael Flanagan, Ian W Davies

ABSTRACT

Chemical modification of siRNA is achieved in a high-throughput manner (96-well plate format) by copper catalyzed azide-alkyne cycloadditions. This transformation can be performed in one synthetic operation at up to four positions with complete specificity, good yield, and acceptable purity. As demonstrated here, this approach extends the current synthetic options for oligonucleotide modifications and simultaneously facilitates the systematic, rapid biological evaluation of modified siRNA.

MATERIALS

Product Number

Brand

Product Description

Supelco

Water, for TOC analysis

Supelco

Water, suitable for ion chromatography

Sigma-Aldrich

Methanol, NMR reference standard

Sigma-Aldrich

Imidazole, ReagentPlus^®, 99%

Sigma-Aldrich

Triethylamine, ≥99.5%

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Methylamine solution, 33 wt. % in absolute ethanol ((denatured with 1% toluene))

Sigma-Aldrich

Methanol, anhydrous, 99.8%

Sigma-Aldrich

2,6-Lutidine, ReagentPlus^®, 98%

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Methylamine solution, 2.0 M in THF

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Methylamine solution, 2.0 M in methanol

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Trichloroacetic acid, BioUltra, ≥99.5% (T)

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Trichloroacetic acid, ACS reagent, for the determination of Fe in blood according to Heilmeyer, ≥99.5%

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Triethylamine, purum, ≥99% (GC)

Sigma-Aldrich

Imidazole, BioUltra, ≥99.5% (GC)

Supelco

Methanol, analytical standard

Sigma-Aldrich

Imidazole, puriss. p.a., ≥99.5% (GC)

Sigma-Aldrich

Azidomethyl phenyl sulfide, 95%

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Methylamine solution, 40 wt. % in H₂O

Supelco

2,6-Lutidine, analytical standard

Sigma-Aldrich

2,6-Lutidine

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Water, for molecular biology, sterile filtered

Sigma-Aldrich

Water, tested according to Ph. Eur.

Sigma-Aldrich

Triethylamine, for protein sequence analysis, ampule, ≥99.5% (GC)

Supelco

Water, for HPCE, for luminescence, suitable for UV/Vis spectroscopy

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Imidazole, BioUltra, for molecular biology, ≥99.5% (GC)

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Water, for cell biology, sterile ultrafiltered

Sigma-Aldrich

Triethylamine, ≥99%

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Water, sterile-filtered, BioReagent, suitable for cell culture

Sigma-Aldrich

Trichloroacetic acid, ≥99.0% (titration)

Sigma-Aldrich

Trichloroacetic acid, suitable for electrophoresis, suitable for fixing solution (for IEF and PAGE gels), ≥99%