DNA mutation detection in a polymer microfluidic network using temperature gradient gel electrophoresis.
DNA mutation detection in a polymer microfluidic network using temperature gradient gel electrophoresis.
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DOI：
10.1021/ac034913y
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发表时间：
2004-02
影响因子：
7.4
通讯作者：
J. Buch;C. Kimball;F. Rosenberger;W. Highsmith;D. DeVoe;Cheng S. Lee
中科院分区：
文献类型：
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作者：
J. Buch;C. Kimball;F. Rosenberger;W. Highsmith;D. DeVoe;Cheng S. Lee
据报道，一种用于 DNA 突变分析的小型化系统，利用聚碳酸酯 (PC) 微流体装置中的温度梯度凝胶电泳 (TGGE)。 TGGE 通过引入热变性梯度来揭示给定异源双链样品中序列异质性的存在，该梯度导致 DNA 序列变体的平均电泳迁移率之间的差异。整体加热器组件被设计并用于沿着分离通道在外部产生空间和时间格式的温度梯度。使用微流体平台中的单通道和 10 通道并行测量来实现模型突变体 DNA 片段的 TGGE 分析，每个片段都包含单碱基替换。此外，还开发并演示了一种包含集成微加热器和传感器阵列的综合聚合物微流体装置，用于执行空间 TGGE 进行 DNA 突变分析。该设备由两个机械粘合在一起的 PC 模块化基板组成。一个基板上压有微通道，另一个基板包含一个锥形微加热器，与一系列温度传感器一起通过光刻图案形成。与外部加热方法相比，集成平台显着降低了功率需求和热响应时间，同时建立了更准确和高效的温度梯度控制，以实现更高的分离分辨率。
A miniaturized system for DNA mutation analysis, utilizing temperature gradient gel electrophoresis (TGGE) in a polycarbonate (PC) microfluidic device, is reported. TGGE reveals the presence of sequence heterogeneity in a given heteroduplex sample by introducing a thermal denaturing gradient that results in differences between the average electrophoretic mobilities of DNA sequence variants. Bulk heater assemblies are designed and employed to externally generate temperature gradients in spatial and temporal formats along the separation channels. TGGE analyses of model mutant DNA fragments, each containing a single base substitution, are achieved using both single- and 10-channel parallel measurements in a microfluidic platform. Additionally, a comprehensive polymer microfluidic device containing an integrated microheater and sensor array is developed and demonstrated for performing spatial TGGE for DNA mutation analysis. The device consists of two PC modular substrates mechanically bonded together. One substrate is embossed with microchannels, and the other contains a tapered microheater, lithographically patterned along with an array of temperature sensors. Compared with the external heating approaches, the integrated platform provides significant reduction in power requirement and thermal response time while establishing more accurate and highly effective control of the temperature gradient for achieving improved separation resolution.