Department of Basic Medical Sciences, Institute of Medical Science, University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639, Japan.
FEBS Open Bio. 2012 Jul 20;2:203-7. doi: 10.1016/j.fob.2012.07.004. Print 2012.
Aptamers are promising gene components that can be used for the construction of synthetic gene circuits. In this study, we isolated an RNA aptamer that specifically inhibits transcription of T7 RNA polymerase (RNAP). The 38-nucleotide aptamer, which was a shortened variant of an initial SELEX isolate, showed moderate inhibitory activity. By stepwise doped-SELEX, we isolated evolved variants with strong inhibitory activity. A 29-nucleotide variant of a doped-SELEX isolate showed 50% inhibitory concentration at 11 nM under typical in vitro transcription conditions. Pull-down experiments revealed that the aptamer inhibited the association of T7 RNAP with T7 promoter DNA.
适体是一种很有前途的基因元件，可用于构建合成基因回路。在这项研究中，我们分离出一种特异性抑制 T7 RNA 聚合酶（RNAP）转录的 RNA 适体。该 38 个核苷酸的适体是初始 SELEX 分离物的缩短变体，显示出中等抑制活性。通过逐步掺杂 SELEX，我们分离出具有强抑制活性的进化变体。掺杂 SELEX 分离物的 29 个核苷酸变体在典型的体外转录条件下，在 11 nM 时表现出 50%的抑制浓度。下拉实验表明，该适体抑制了 T7 RNAP 与 T7 启动子 DNA 的结合。
