Rapid prototyping of multichannel microfluidic devices for single-molecule DNA curtain imaging.
Rapid prototyping of multichannel microfluidic devices for single-molecule DNA curtain imaging.
复制标题
DOI：
10.1021/ac500267v
复制
发表时间：
2014-04
影响因子：
7.4
通讯作者：
A. D. Robison;Ilya J. Finkelstein
中科院分区：
文献类型：
--
作者：
A. D. Robison;Ilya J. Finkelstein
单分子成像和生化反应的操纵继续揭示许多生物学见解。为了促进这些研究，我们已经开发并实施了一种高通量的方法来组织和成像数百个单独的DNA分子在对齐的扩散屏障。尽管如此，在各种反应条件下获得统计相关数据集仍然具有挑战性。在这里，我们提出了一种集成高通量单分子“DNA幕”成像与聚（二甲基硅氧烷）（PDMS）为基础的微流体的方法。我们的台式制造方法可以在几分钟内完成，使用所有分子生物学实验室中的常用工具。我们证明了这种方法的实用性，通过同时成像的两个独立的生化反应条件下的层流装置。此外，在被动线性梯度发生器中可以同时观察到五种不同的反应条件。将快速微流体制造与高通量DNA幕相结合，极大地扩展了我们询问复杂生物反应的能力。
Single-molecule imaging and manipulation of biochemical reactions continues to reveal numerous biological insights. To facilitate these studies, we have developed and implemented a high-throughput approach to organize and image hundreds of individual DNA molecules at aligned diffusion barriers. Nonetheless, obtaining statistically relevant data sets under a variety of reaction conditions remains challenging. Here, we present a method for integrating high-throughput single-molecule "DNA curtain" imaging with poly(dimethylsiloxane) (PDMS)-based microfluidics. Our benchtop fabrication method can be accomplished in minutes with common tools found in all molecular biology laboratories. We demonstrate the utility of this approach by simultaneous imaging of two independent biochemical reaction conditions in a laminar flow device. In addition, five different reaction conditions can be observed concurrently in a passive linear gradient generator. Combining rapid microfluidic fabrication with high-throughput DNA curtains greatly expands our capability to interrogate complex biological reactions.