The synthesis for the phosphoramidite of 8-oxoG,[19 (link)] 8-oxoA,[31 (link)] and 8-oxoI/8-BrI[32 (link)] were previously reported by us and the same methodology was used to prepare all oligonucleotides, via solid-phase synthesis. The synthesis of oligonucleotides of RNA containing 8-BrA was not possible in our hands due to its transformation to the corresponding 8-methylamine derivative (upon AMA-deprotection of the synthesized oligonucleotide, Figure S-4 ). It is possible that oligonucleotides containing this modified nucleoside can be attained by varying the deprotection conditions, however we have not been yet successful. UV-vis spectroscopy of all small molecules was carried out on a Perkin Elmer λ−650 UV/vis spectrometer using quartz cuvettes (1 cm pathlength). All experiments described herein were carried out in triplicate.
λ 650 uv vis spectrometer
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The λ-650 UV/vis spectrometer is a laboratory instrument designed to measure the absorbance or transmittance of light in the ultraviolet and visible regions of the electromagnetic spectrum. It is capable of analyzing the spectral characteristics of samples.
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4 protocols using λ 650 uv vis spectrometer
1
Synthesis of Modified Nucleoside Phosphoramidites
2
UV/Vis Spectroscopic Characterization of Oligonucleotides
3
Phosphoramidite Synthesis of Modified Nucleosides
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The detailed synthesis for the phosphoramidites of 8-oxoI and 8-BrI is included in the (S1 and S2 Files). It includes spectroscopic information for all known and novel intermediates. 1H NMR, 13C NMR, and 31P NMR spectra, recorded at 300, 75, and 121.5 MHz respectively (using a standard broadband multinuclear probe on a 300MHz Avance III platform from Bruker); IR spectra were recorded on a diamond ATR sampler using powders of pure materials; high-resolution mass spectrometry was carried out via ESI/APCI; and UV-vis spectroscopy of all small molecules was carried out on a Perkin Elmer λ-650 UV/vis spectrometer using quartz cuvettes (1 cm pathlength). The synthesis corresponding to the phosphoramidite for 8-oxoG was carried out according to a previous report [40 (link)]. All experiments described herein were carried out in triplicate, unless otherwise noted. The protocol for the reverse transcription experiments is available at: dx.doi.org/10.17504/protocols.io.birskd6e .
4
Synthesis of Modified Nucleoside Phosphoramidites
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The synthesis for the phosphoramidite of 8‐oxoG,[19 ] 8‐oxoA,[31 ] and 8‐oxoI/8‐BrI[32 ] were previously reported by us and the same methodology was used to prepare all oligonucleotides, via solid‐phase synthesis. The synthesis of oligonucleotides of RNA containing 8‐BrA was not possible in our hands due to its transformation to the corresponding 8‐methylamine derivative (upon AMA‐deprotection of the synthesized oligonucleotide, Figure S‐4 ). It is possible that oligonucleotides containing this modified nucleoside can be attained by varying the deprotection conditions, however we have not been yet successful. UV‐vis spectroscopy of all small molecules was carried out on a Perkin Elmer λ‐650 UV/vis spectrometer using quartz cuvettes (1 cm pathlength). All experiments described herein were carried out in triplicate.
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