School of Environmental Science and Technology, Key Laboratory of Industrial Ecology and Environmental Engineering, (Ministry of Education), Dalian University of Technology, Dalian, 116024, China.
Institute of Environmental Science, Shanxi University, Taiyuan, 030006, China.
Angew Chem Int Ed Engl. 2020 Dec 14;59(51):22947-22951. doi: 10.1002/anie.202010693. Epub 2020 Nov 9.
φ29 DNA polymerase (Polφ29) is capable of synthesizing long-chain single-stranded (ss) DNA molecules by copying the sequence of a small ss circular DNA template (ssCDT) in a process known as rolling circle amplification (RCA). The use of a ssCDT in RCA, however, comes with a key drawback: the rate of DNA synthesis is significantly reduced. We hypothesize that this issue can be overcome using a very long linear ssDNA template with a repeating sequence. To test this idea, we engineered a DNA assembly, which we denote "micrometer-sized DNA track" (μDT). This μDT, with an average length of ≈13.5 μm, is made of a long chain DNA with a primer-binding domain at its 3' end and ≈1000 repeating sequence units at its 5' end, each carrying a DNA anchor. We find that Polφ29 copies μDT at a speed ≈5-time faster than it does a related ssCDT. We use this to design a simple all-in-one printed paper device for rapid and sensitive detection of microRNA let-7. This paper sensor is capable of detecting 1 pM let-7a in 10 minutes.
φ29 DNA 聚合酶（Polφ29）能够通过复制小 ss 圆形 DNA 模板（ssCDT）的序列来合成长链单链（ss）DNA 分子，这一过程称为滚环扩增（RCA）。然而，在 RCA 中使用 ssCDT 存在一个关键的缺点：DNA 合成的速度显著降低。我们假设，使用具有重复序列的非常长的线性 ssDNA 模板可以克服这个问题。为了验证这个想法，我们设计了一种 DNA 组装体，我们称之为“微米级 DNA 轨道”（μDT）。这种 μDT 的平均长度约为 13.5μm，由一条长链 DNA 组成，其 3' 端有一个引物结合域，5' 端有大约 1000 个重复序列单元，每个单元都带有一个 DNA 锚。我们发现，Polφ29 复制 μDT 的速度比复制相关的 ssCDT 快约 5 倍。我们利用这一点设计了一种简单的一体化印刷纸质设备，用于快速灵敏地检测 microRNA let-7。这种纸传感器能够在 10 分钟内检测到 1pM 的 let-7a。
