Synthesis of azobenzene-tethered DNA for reversible photo-regulation of DNA functions: hybridization and transcription
Synthesis of azobenzene-tethered DNA for reversible photo-regulation of DNA functions: hybridization and transcription
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DOI：
10.1038/nprot.2006.465
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发表时间：
2007-01-01
期刊：
影响因子：
14.8
通讯作者：
Komiyama, Makoto
中科院分区：
文献类型：
--
作者：
Asanuma, Hiroyuki;Liang, Xingguo;Komiyama, Makoto
由D-苏氨醇合成带有偶氮苯的亚磷酰胺单体。使用该单体，偶氮苯部分可以在常规DNA合成仪上的任何位置引入寡脱氧核糖核苷酸（DNA）中。有了这种偶氮苯拴系的DNA，DNA双链体的形成和解离可以通过偶氮苯的顺-反异构化可逆地进行光调控。当偶氮苯采取反式形式时，形成稳定的双链体。在通过UV光照射（300 nm < lambda < 400 nm）将反式偶氮苯异构化成其顺式形式后，双链体可以解离成两条链。双链体在光诱导的顺反异构化（λ> 400 nm）上重新形成。在T7启动子的特定位置引入偶氮苯也有效地和可逆地通过T7-RNA聚合酶光调节转录。可逆调节可以重复多次，而不会对DNA或偶氮苯部分造成损伤。这些程序大约需要10天才能完成。
A phosphoramidite monomer bearing an azobenzene is synthesized from D-threoninol. Using this monomer, azobenzene moieties can be introduced into oligodeoxyribonucleotide (DNA) at any position on a conventional DNA synthesizer. With this azobenzene-tethered DNA, formation and dissociation of a DNA duplex can be reversibly photo-regulated by cis-trans isomerization of the azobenzene. When the azobenzene takes a trans-form, a stable duplex is formed. After isomerization of the trans-azobenzene to its cis-form by UV-light irradiation (300 nm < lambda < 400 nm), the duplex can be dissociated into two strands. The duplex is reformed on photo-induced cis-trans isomerization (lambda > 400 nm). The introduction of azobenzenes into the T7 promoter at specific positions also efficiently and reversibly photo-regulates transcription by T7-RNA polymerase. The reversible regulation can be repeated many times without causing damage to the DNA or the azobenzene moiety. These procedures take approximately 10 d to complete.