Center for Nano and Micro Mechanics, School of Aerospace Engineering, Tsinghua University, Beijing, People's Republic of China.
Nanotechnology. 2019 Jan 25;30(4):045101. doi: 10.1088/1361-6528/aaeddc.
DNA combing is a widely used method for stretching and immobilising DNA molecules on a surface. Fluorescent labelling of genomic information enables high-resolution optical analysis of DNA at the single-molecule level. Despite its simplicity, the application of DNA combing in diagnostic workflows is still limited, mainly due to difficulties in analysing multiple small-volume DNA samples in parallel. Here, we report a simple and versatile microfluidic DNA combing technology (μDC), which allows manipulating, stretching and imaging of multiple, microliter scale DNA samples by employing a manifold of parallel microfluidic channels. Using DNA molecules with repetitive units as molecular rulers, we demonstrate that the μDC technology allows uniform stretching of DNA molecules. The stretching ratio remains consistent along individual molecules as well as between different molecules in the various channels, allowing simultaneous quantitative analysis of different samples loaded into parallel channels. Furthermore, we demonstrate the application of μDC to characterise UVB-induced DNA damage levels in human embryonic kidney cells and the spatial correlation between DNA damage sites. Our results point out the potential application of μDC for quantitative and comparative single-molecule studies of genomic features. The extremely simple design of μDC makes it suitable for integration into other microfluidic platforms to facilitate high-throughput DNA analysis in biological research and medical point-of-care applications.
DNA 梳状技术是一种广泛用于拉伸和固定 DNA 分子在表面上的方法。对基因组信息进行荧光标记可以实现单个分子水平上的高分辨率光学分析 DNA。尽管它很简单，但 DNA 梳状技术在诊断工作流程中的应用仍然受到限制，主要是因为难以平行分析多个小体积的 DNA 样本。在这里，我们报告了一种简单而通用的微流控 DNA 梳状技术（μDC），该技术允许通过采用多个平行微流道来操纵、拉伸和成像多个微升规模的 DNA 样本。使用具有重复单元的 DNA 分子作为分子标尺，我们证明 μDC 技术允许 DNA 分子均匀拉伸。在单个分子以及各个通道中的不同分子之间，拉伸比保持一致，允许同时对加载到平行通道中的不同样本进行定量分析。此外，我们还展示了 μDC 在分析人类胚胎肾细胞中 UVB 诱导的 DNA 损伤水平以及 DNA 损伤部位之间的空间相关性方面的应用。我们的结果指出了 μDC 在定量和比较基因组特征的单分子研究中的潜在应用。μDC 的极其简单的设计使其适合集成到其他微流控平台中，以促进生物研究和医疗即时检测应用中的高通量 DNA 分析。
