The fluorescent adenine analogue qAN4 was recently shown to possess promising photophysical properties, including a high brightness as a monomer. Here we report the synthesis of the phosphoramidite of qAN4 and its successful incorporation into DNA oligonucleotides using standard solid-phase synthesis. Circular dichroism and thermal melting studies indicate that the qAN4-modification has a stabilizing effect on the B-form of DNA. Moreover, qAN4 base-pairs selectively with thymine with mismatch penalties similar to those of mismatches of adenine. The low energy absorption band of qAN4 inside DNA has its peak around 358 nm and the emission in duplex DNA is partly quenched and blue-shifted (ca. 410 nm), compared to the monomeric form. The spectral properties of the fluorophore also show sensitivity to pH; a property that may find biological applications. Quantum yields in single-stranded DNA range from 1-29 % and in duplex DNA from 1-7 %. In combination with the absorptive properties, this gives an average brightness inside duplex DNA of 275 M-1  cm-1 , more than five times higher than the most used environment-sensitive fluorescent base analogue, 2-aminopurine. Finally, we show that qAN4 can be used to advantage as a donor for interbase FRET applications in combination with adenine analogue qAnitro as an acceptor.

中文翻译：

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用对环境敏感的亮腺嘌呤类似物qAN4点亮DNA。

最近显示，荧光腺嘌呤类似物qAN4具有令人满意的光物理特性，包括作为单体的高亮度。在这里，我们报告qAN4的亚磷酰胺的合成及其使用标准固相合成法成功地掺入DNA寡核苷酸的过程。圆二色性和热解链研究表明，qAN4修饰对DNA的B型具有稳定作用。而且，qAN4碱基对与胸腺嘧啶选择性地具有错配罚分，类似于腺嘌呤的错配。与单体形式相比，DNA内部qAN4的低能吸收带具有约358 nm的峰值，双链体DNA中的发射被部分淬灭和蓝移（约410 nm）。荧光团的光谱特性也显示出对pH的敏感性。可能找到生物学用途的特性。单链DNA的量子产率为1-29％，双链DNA的量子产率为1-7％。结合吸收特性，双链DNA内部平均亮度为275 M-1 cm-1，比最常用的对环境敏感的荧光碱基类似物2-氨基嘌呤高出五倍以上。最后，我们证明qAN4可以与腺嘌呤类似物qAnitro作为受体结合使用，作为碱基间FRET应用的供体。