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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, suitable for ion chromatography

Imidazole, European Pharmacopoeia (EP) Reference Standard

Sigma-Aldrich

Water, BioPerformance Certified

Supelco

Density Standard 998 kg/m³, H&D Fitzgerald Ltd. Quality

Supelco

Water, ACS reagent, for ultratrace analysis

Sigma-Aldrich

Benzyl azide solution, ~0.5 M in dichloromethane, ≥95.0% (HPLC)

Millipore

Acetonitrile solution, suitable for HPLC, acetonitrile:water 5:95% (v/v), 10 mM Ammoniumbicarbonate, pH 10,0

Sigma-Aldrich

Methanol, anhydrous, 99.8%

Sigma-Aldrich

Water, deuterium-depleted, ≤1 ppm (Deuterium oxide)

Supelco

2,6-Lutidine, analytical standard

Sigma-Aldrich

Water, for cell biology, sterile ultrafiltered

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Azidomethyl phenyl sulfide, 95%

Sigma-Aldrich

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

Supelco

Methanol, analytical standard

Supelco

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

Sigma-Aldrich

Triethylamine, BioUltra, ≥99.5% (GC)

Sigma-Aldrich

Water, tested according to Ph. Eur.

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Water, for molecular biology, sterile filtered

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

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Imidazole, ReagentPlus^®, 99%

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

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Methylamine solution, 2.0 M in THF

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2,6-Lutidine, ReagentPlus^®, 98%

Sigma-Aldrich

2,6-Lutidine

Sigma-Aldrich

Imidazole, for molecular biology, ≥99% (titration)

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Imidazole, ≥99% (titration), crystalline

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

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Trichloroacetic acid, BioXtra, ≥99.0%

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Water, for embryo transfer, sterile-filtered, BioXtra, suitable for mouse embryo cell culture

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Water, PCR Reagent