A binary Cy3 aptamer probe composed of folded modules
A binary Cy3 aptamer probe composed of folded modules
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DOI：
10.1016/j.ab.2010.01.015
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发表时间：
2010-05-01
影响因子：
2.9
通讯作者：
Nakamura, Yoshikazu
中科院分区：
文献类型：
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作者：
Endo, Kei;Nakamura, Yoshikazu
适体是短的单链DNA或RNA序列，其基于其对靶分子的高亲和力在体外选择。染料结合适体是一种很有前途的工具，不仅可以实时检测DNA或RNA序列，还可以在体外和体内检测感兴趣的蛋白质。在这项研究中，我们的目的是分离的RNA适体Cy 3，一个广泛使用的，膜渗透，无毒的荧光花青染料。对Cy 3的亲和RNA分子的广泛选择产生了命名为Cy3_apt的独特序列适体。所选择的Cy3_apt为83个核苷酸长，并成功缩短至49个核苷酸长，通过多个碱基变化增加了对Cy 3的亲和力。最短的Cy3_apt由两个单独的发夹模块组成，所述发夹模块是通过表面等离子体共振（SPR）测定监测的对Cy 3的亲和力所需的。此外，Cy 3的荧光在与Cy3_apt结合时增加。Cy3_apt的两个模块在彼此分离时充当二元适体探针。我们证明了二元Cy3_apt探针适用于通过标记靶序列来检测靶寡核苷酸或RNA-RNA相互作用。该二元探针由两个折叠模块组成，称为折叠二元探针。(C)2010年爱思唯尔公司所有的紧身衣保留。
Aptamers are short single-stranded DNA or RNA sequences that are selected in vitro based on their high affinity to a target molecule. Dye-binding aptamers are promising tools for real-time detection of not only DNA or RNA sequences but also proteins of interest both in vitro and in vivo. In this study, we aimed to isolate an RNA aptamer to Cy3, a widely used, membrane-permeant, and nontoxic fluorescent cyanine dye. Extensive selection of affinity RNA molecules to Cy3 yielded a unique sequence aptamer named Cy3_apt. The selected Cy3_apt was 83 nucleotides long and successfully shortened to 49 nucleotides long with increased affinity to Cy3 by multiple base changes. The shortest Cy3_apt is composed of two separate hairpin modules that are required for the affinity to Cy3 as monitored by the surface plasmon resonance (SPR) assay. Also, the fluorescence of Cy3 increased on binding to Cy3_apt. The two modules of Cy3_apt, when detached from each other, functioned as a binary aptamer probe. We demonstrate that the binary Cy3_apt probe is applicable to the detection of target oligonucleotides or RNA-RNA interaction by tagging with target sequences. This binary probe consists of two folded modules, referred to as a folded binary probe. (C) 2010 Elsevier Inc. All tights reserved.