Separation of long DNA molecules by quartz nanopillar chips under a direct current electric field
Separation of long DNA molecules by quartz nanopillar chips under a direct current electric field
复制标题
DOI：
10.1021/ac030303m
复制
发表时间：
2004-01-01
影响因子：
7.4
通讯作者：
Baba, Y
中科院分区：
文献类型：
--
作者：
Kaji, N;Tezuka, Y;Baba, Y
我们已经建立了在石英芯片微通道内构建高纵横比（直径100- 500nm，高500- 5000nm）纳米柱的纳米制造技术。柱的大小和柱之间的间距被设计为DNA筛分矩阵，用于在几千碱基对（kbp）以上的大DNA片段的最佳分析。基于优化设计，研制了纳米柱通道和简单交叉进样器芯片，应用于常规凝胶电泳和毛细管电泳难以分离的DNA片段（1-38 kbp）和大片段（lambda DNA, 48.5 kbp; T4 DNA, 165.6 kbp）的分离。在1 ~ 38kbp范围内的DNA片段被分离成清晰的条带，并且在直流电场下，λ DNA和T4 DNA的混合物通过380 μ m长的纳米柱通道在10 s内成功分离。通道（380-1450微米长）的理论板数N为1000-3000 (0.7 × 10（6)-2.1 × 10(6)板/m）。在纳米柱通道中对单个DNA分子的电泳观察表明，最优的纳米柱诱导T4 DNA在电泳过程中形成窄的u形构象，而λ DNA在电泳过程中保持较球形的构象。我们证明，即使在直流电场下，最佳的纳米柱尺寸取决于DNA分子的旋转半径，这使得在短时间内分离大的DNA片段成为可能。
We have established the nanofabrication technique for constructing nanopillars with high aspect ratio (100-500 nm diameter and 500-5000 nm tall) inside a microchannel on a quartz chip. The size of pillars and the spacing between pillars are designed as a DNA sieving matrix for optimal analysis of large DNA fragments over a few kilobase pairs (kbp). A chip with nanopillar channel and simple cross injector was developed based on the optimal design and applied to the separation of DNA fragments (1-38 kbp) and large DNA fragments (lambda DNA, 48.5 kbp; T4 DNA, 165.6 kbp) that are difficult to separate on conventional gel electrophoresis and capillary electrophoresis without a pulsed-field technique. DNA fragments ranging from 1 to 38 kbp were separated as clear bands, and furthermore, the mixture of lambda DNA and T4 DNA was successfully separated by a 380-mum-long nanopillar channel within only 10 s even under a direct current (dc) electric field. Theoretical plate number N of the channel (380-1450 mum long) was 1000-3000 (0.7 x 10(6)-2.1 x 10(6) plates/m). A single DNA molecule observation during electrophoresis in a nanopillar channel revealed that the optimal nanopillars induced T4 DNA to form a narrow U-shaped conformation during electrophoresis whereas lambda DNA kept a rather spherical conformation. We demonstrated that, even under a dc electric field, the optimal nanopillar dimensions depend on a gyration radius of DNA molecule that made it possible to separate large DNA fragments in a short time.