Yanfei Zhang, Yue Mou, Meiyun Chen, Xinru Lin, Yujie Zhao and Xingyu Luo
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引用次数: 0
摘要
乙型肝炎病毒(HBV)是一种严重威胁公共健康的病毒,可导致慢性肝病。虽然可以通过估计 HBV DNA 基因组的容量来量化评估 HBV 感染情况,但仍然缺乏一种不使用酶或化学标记的有效而稳健的检测方法。在此,我们设计了一种二元分裂荧光 DNA 合感器(bsFDA),它通过合理地将莴苣合感器分裂成两条功能性 DNA 短链,并利用 HBV DNA 片段互补序列(HDs)。在这一策略中,研究人员研究了bsFDA对HDs的特异性识别能力,与莴苣适配体结构形成三重DNA。同时,当 DFHO 与三重 DNA 结构形成荧光复合物时,bsFDA 就会发出开启荧光,从而实现无酶、无标记、快速反应和可靠的荧光读数,用于检测 HDs 和潜在的 HDs 突变体。此外,bsFDA 还被用于分析不同真实基质(包括人血清和细胞裂解液)中的加标 HDs。bsFDA令人满意的回收率和重现性揭示了它在生物样品中进行 HDs 分析的潜在检测功效。总之,bsFDA 在开发功能化相应传感器和实现生物研究中的病毒基因组分析方面具有巨大潜力。
Binary split fluorescent biosensor based on lettuce DNA aptamer for label-free and enzyme-free analysis of hepatitis B viral DNA†
Hepatitis B virus (HBV) acts as a severe public health threat, causing chronic liver diseases. Although the quantified evaluation of HBV infection can be obtained by estimating the capacity of the HBV DNA genome, it still lacks an effective and robust detection method without using enzymes or chemical labeling. Herein, we have designed a binary split fluorescent DNA aptasensor (bsFDA) by rationally splitting the lettuce aptamer into two functional DNA short chains and utilizing the HBV DNA segment complementary sequences (HDs). In this strategy, the bsFDA has been investigated to specifically recognize the HDs, forming a triplex DNA with the lettuce aptamer structure. Meanwhile, the turn-on fluorescence of bsFDA is obtained upon formation of a fluorescent complex between DFHO and the triplex DNA structure, allowing the enzyme-free, label-free, fast-responsive, and reliable fluorescence readout for detecting HDs and the potential HDs mutants. Moreover, bsFDA has been applied for spiked HDs analysis in different real matrixes, including human serum and cell lysate. The satisfactory recovery rates and reproducibility of the bsFDA reveal its potential detection efficacy for HDs analysis in biological samples. Overall, bsFDA holds great potential in developing functionalized aptasensors and realizing viral genome analysis in biological research.