Lambda25

HPLC analysis was carried out using an Agilent 1100 series system coupled with a photodiode array detector. UV and FT-IR spectra were obtained employing a Perkin Elmer Lambda25 and a Jasco FT/IR-4200 instrument, respectively. All NMR spectra were recorded on Bruker Avance III 400 and 600 spectrometers. Spectra were referenced to the residual solvent signal of acetone-d₆ with resonances at δ_H/C 2.04/29.8. HR-ESI-TOF-MS data were recorded on a Bruker maXis 4G mass spectrometer. ESI-LC/MS experiments were carried out using an Agilent 1200 series system coupled with an ESI spectrometer (LC/MSD Ultra Trap System XCT 6330).

The spectral measurements were made with a UV-Vis Perkin-Elmer Lambda25 and an FTIR Jasco 6300 spectrometer.
An ATR accessory equipped with a diamond crystal (Pike Technologies, Madison, Wisconsin, USA) allows the collection of FTIR spectra directly on a sample without any special preparation. The FTIR spectra were recorded in the region of 4000–400 cm⁻¹, with a TGS detector, and apodization Cosine. The spectral data were processed with JASCO Spectra Manager software, version 2. Samples were scanned at a 4 cm⁻¹ resolution, accumulation: 100 scans. Background reference spectra were recorded using air after every sample to minimize the interference due to carbon dioxide and water vapor in the atmosphere. Between measurements, the ATR crystal was carefully cleaned using pure acetone (Sigma-Aldrich Co., Saint Louis, MO, USA), then, it was dried with soft tissue [43 (link),44 (link),45 (link)].
All measurements were taken at room temperature (T = 23 °C). For each sample, three replicate spectra were recorded to ensure spectral reproducibility and to assess analytical precision; then, the average spectrum was complete.

Unlabeled and labeled oligonucleotides were prepared in 50 mM Tris–HCl, pH 7.5, with appropriate salt as stated in the text, and had a final DNA concentration of 2.5 μM. UV or CD spectra were obtained using a UV/Vis spectrophotometer (PerkinElmer Lambda 25) or a CD spectrophotometer (Jasco J-810) measuring the sample in a 1 cm path length 3 ml or 100 μl quartz cuvette (Hellma), respectively. All melting curves were acquired at 295 nm with a gradient of 0.2°C min⁻¹. Melting curves were plotted as described in ref. (66 (link)). CD spectra were recorded at room temperature unless otherwise specified and background corrected before analysis.