A DNA prism for high-speed continuous fractionation of large DNA molecules
A DNA prism for high-speed continuous fractionation of large DNA molecules
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DOI：
10.1038/nbt733
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发表时间：
2002-10-01
影响因子：
46.9
通讯作者：
Cox, EC
中科院分区：
文献类型：
--
作者：
Huang, LR;Tegenfeldt, JO;Cox, EC
大DNA分子的分析和分离在许多基因组计划中起着关键作用。标准方法脉冲场凝胶电泳法(PFGE)速度很慢，运行时间从10小时到200多小时不等。在这份报告中，我们描述了一个缩略图大小的设备，它可以在15秒内对大的DNA片段(61-209千碱基(Kb))进行分类，分辨率接近13%。一组微米级的柱子用作筛分基质，集成的微流控通道在空间上塑造基质上的电场。非对称脉冲场被应用于连续流动操作，根据DNA分子的分子质量在不同方向上进行分类，就像棱镜在不同角度偏转不同波长的光一样。我们通过使用该设备分离从细菌人工染色体制备的大型DNA插入物来证明该设备的稳健性，细菌人工染色体是大多数基因组学项目广泛使用的DNA来源。
The analysis and fractionation of large DNA molecules plays a key role in many genome projects. The standard method, pulsed-field gel electrophoresis (PFGE), is slow, with running times ranging from 10 hours to more than 200 hours. In this report, we describe a thumbnail-sized device that sorts large DNA fragments (61-209 kilobases (kb)) in 15 seconds, with a resolution of similar to13%. An array of micron-scale posts serves as the sieving matrix, and integrated microfluidic channels spatially shape the electric fields over the matrix. Asymmetric pulsed fields are applied for continuous-flow operation, which sorts DNA molecules in different directions according to their molecular masses, much as a prism deflects light of different wavelengths at different angles. We demonstrate the robustness of the device by using it to separate large DNA inserts prepared from bacterial artificial chromosomes, a widely used DNA source for most genomics projects.