College of Chemistry and Chemical Engineering, State Key Laboratory of Physical Chemistry of Solid Surfaces, Xiamen University, Xiamen 361005, China.
Biosens Bioelectron. 2011 Jan 15;26(5):2796-800. doi: 10.1016/j.bios.2010.11.005. Epub 2010 Nov 16.
Rapid growth of available sequence data has made the detection of nucleic acids critical to the development of modern life sciences. Many amplification methods based on gold nanoparticles and endonuclease for sensitive DNA detection have been developed. However, these approaches require specific target sequence for endonuclease recognition, which cannot be fulfilled in all systems. Replacing the restriction enzyme with a nuclease that does not require any specific recognition sequence may offer a universally adaptable system. Here we have developed a novel homogeneous, colorimetric DNA detection method, which consists of Exo III, a linker DNA, and two DNA-modified gold nanoparticles. This system is simple, low-cost, sensitive and selective. By coupling cyclic enzymatic cleavage and gold nanoparticle for signal amplification, our system provides a colorimetric detection limit of 15 pM, which is 3 orders of magnitude more sensitive than that of a general three-component sandwich assay format. Due to the intrinsic property of Exo III, our method shows excellent detection selectivity for single-base discrimination. More importantly, superior to other methods based on nicking and FokI endonuclease, our target sequence-independent platform is generally applicable for DNA sensing. This new approach could be widely applied to sensitive nucleic acids detection.
大量可用序列数据的快速增长使得核酸检测对于现代生命科学的发展至关重要。已经开发出了许多基于金纳米颗粒和内切酶的用于灵敏 DNA 检测的扩增方法。然而，这些方法需要针对内切酶识别的特定靶序列，而这在所有系统中都无法满足。用不需要任何特定识别序列的核酸酶替代限制性内切酶可能会提供一个普遍适用的系统。在这里，我们开发了一种新颖的均相、比色 DNA 检测方法，该方法由 Exo III、连接 DNA 和两个 DNA 修饰的金纳米颗粒组成。该系统简单、低成本、灵敏且具有选择性。通过偶联循环酶切和金纳米颗粒进行信号放大，我们的系统提供了 15 pM 的比色检测限，比一般的三组分夹心测定格式灵敏 3 个数量级。由于 Exo III 的固有特性，我们的方法对单碱基鉴别具有出色的检测选择性。更重要的是，与基于切口和 FokI 内切酶的其他方法相比，我们的无靶序列平台通常适用于 DNA 传感。这种新方法可以广泛应用于灵敏的核酸检测。
