Herein, we report a new probe for the determination of the concentration of NF-κB p50, one kind of DNA-binding transcription factors (TFs), by using Exonuclease III (Exo III)-aided amplification and gold nanoparticle mediated fluorescence intensity. Since TFs play critical roles in various biological processes, the detection of TFs can provide a lot of useful biological information for studding gene expression regulation related disease. In our system, in the presence of transcription factor, Exo III based amplification reaction was trigged. This enzymatic digestion results in the release of intermediate DNA and ultimately liberating the fluorophore (which, separated from the quencher of AuNP and BHQ2, now fluoresces). The released intermediate DNA then hybridizes with another strand3, whence the cycle starts anew. So, the fluorescence intensity reflects the NF-κB p50 concentration with a detection limit of 1.32 pM. Importantly, this method might be further extended to selectively detect various dsDNA-binding proteins by simply changing the binding-site sequences of strand1/strand2 duplex probes.

中文翻译：

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利用核酸外切酶III辅助扩增和金纳米粒子介导的荧光强度测定转录因子浓度：一种基因转录相关酶检测的新方法

在此，我们报告了一种新探针，用于通过使用核酸外切酶 III (Exo III) 辅助扩增和金纳米颗粒介导的荧光强度来测定 NF-κB p50，一种 DNA 结合转录因子 (TF) 的浓度。由于TFs在各种生物过程中起着关键作用，TFs的检测可以为研究基因表达调控相关疾病提供大量有用的生物学信息。在我们的系统中，在转录因子存在的情况下，触发了基于 Exo III 的扩增反应。这种酶消化导致中间体 DNA 的释放并最终释放荧光团（它与 AuNP 和 BHQ2 的猝灭剂分离，现在发出荧光）。然后释放的中间 DNA 与另一条链 3 杂交，从而重新开始循环。所以，荧光强度反映了 NF-κB p50 浓度，检测限为 1.32 pM。重要的是，这种方法可能会进一步扩展到通过简单地改变链 1/链 2 双链探针的结合位点序列来选择性检测各种 dsDNA 结合蛋白。