Zhu Longjiao, Shao Xiangli, Luo Yunbo, Huang Kunlung, Xu Wentao
Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Food Science & Nutritional Engineering, China Agricultural University , Beijing 100083, China.
Beijing Laboratory for Food Quality and Safety, College of Food Science & Nutritional Engineering, China Agricultural University , Beijing 100083, China.
ACS Chem Biol. 2017 May 19;12(5):1373-1380. doi: 10.1021/acschembio.7b00060. Epub 2017 Apr 4.
A two-way colorimetric biosensor based on unmodified gold nanoparticles (GNPs) and a switchable double-stranded DNA (dsDNA) concatemer have been demonstrated. Two hairpin probes (H1 and H2) were first designed that provided the fuels to assemble the dsDNA concatemers via hybridization chain reaction (HCR). A functional hairpin (FH) was rationally designed to recognize the target sequences. All the hairpins contained a single-stranded DNA (ssDNA) loop and sticky end to prevent GNPs from salt-induced aggregation. In the presence of target sequence, the capture probe blocked in the FH recognizes the target to form a duplex DNA, which causes the release of the initiator probe by FH conformational change. This process then starts the alternate-opening of H1 and H2 through HCR, and dsDNA concatemers grow from the target sequence. As a result, unmodified GNPs undergo salt-induced aggregation because the formed dsDNA concatemers are stiffer and provide less stabilization. A light purple-to-blue color variation was observed in the bulk solution, termed the light-off sensing way. Furthermore, H1 ingeniously inserted an aptamer sequence to generate dsDNA concatemers with multiple small molecule binding sites. In the presence of small molecule targets, concatemers can be disassembled into mixtures with ssDNA sticky ends. A blue-to-purple reverse color variation was observed due to the regeneration of the ssDNA, termed the light-on way. The two-way biosensor can detect both nucleic acids and small molecule targets with one sensing device. This switchable sensing element is label-free, enzyme-free, and sophisticated-instrumentation-free. The detection limits of both targets were below nanomolar.
基于未修饰的金纳米颗粒（GNP）和可切换双链DNA（dsDNA）串联体的双向比色生物传感器已得到证实。首先设计了两种发夹探针（H1和H2），它们作为燃料通过杂交链式反应（HCR）组装dsDNA串联体。合理设计了一种功能性发夹（FH）以识别靶序列。所有发夹都包含一个单链DNA（ssDNA）环和粘性末端，以防止GNP因盐诱导而聚集。在存在靶序列的情况下，FH中被阻断的捕获探针识别靶标形成双链DNA，这会导致FH构象变化从而释放引发探针。此过程随后通过HCR启动H1和H2的交替打开，dsDNA串联体从靶序列开始生长。结果，未修饰的GNP会发生盐诱导聚集，因为形成的dsDNA串联体更硬且提供的稳定性更低。在本体溶液中观察到从浅紫色到蓝色的颜色变化，称为关闭型传感方式。此外，H1巧妙地插入了一个适体序列，以生成具有多个小分子结合位点的dsDNA串联体。在存在小分子靶标的情况下，串联体可分解为具有ssDNA粘性末端的混合物。由于ssDNA的再生，观察到从蓝色到紫色的反向颜色变化，称为开启型方式。这种双向生物传感器可以用一个传感装置检测核酸和小分子靶标。这种可切换的传感元件无需标记、无需酶且无需复杂仪器。两种靶标的检测限均低于纳摩尔。
