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Fields & Applications Genomics & DNA Analysis

Go With the Flow

The problem

Standard gel electrophoresis has been intensively used for DNA fractionation in various genotyping and sequencing applications, and has the advantages of great simplicity, versatility, and reproducibility. However, it suffers from long processing times – many hours or even days; for instance, a typical pulsed-field gel electrophoresis (PFGE) device takes 25 hours to perform fractionation of 5–120 kbp DNA. Given trends towards second-generation sequencing, replacing standard gel electrophoresis with microchip-based systems is an attractive option to minimize processing time and optimize DNA fractionation. We set out to develop a new method to improve sample throughput and recovery compared with current technology.

Background

The development of DNA electrophoresis, using both slab gels and capillaries, enabled the first-generation DNA sequencing and genotyping technologies required for the Human Genome Project. However, only minimal sample numbers could be separated and identified, at very high cost. In recent years, efforts have been directed towards reducing the cost and analysis time of DNA genotyping in second- and later-generation sequencing tools. To achieve this, sample preparation methods, such as electrophoresis, must be optimized to increase the throughput and efficiency of the analyses via user-friendly, portable and functional platforms. In some devices, traditional DNA separation gels have been replaced by microfabricated post arrays for separation of genomic-length DNA. However, defect-free fabrication of 3D nanostructures, such as nanopost arrays and crystalline nanoarrays, is extremely challenging. 2D nanostructures are easier to manufacture, but intrinsically yield low sample throughput. An ideal sieving matrix should have simple design and fabrication steps, yet provide high-resolution, high-throughput separation. And that’s why we opted for gel-based devices. People had originally considered continuous flow separation in such devices impossible, but we showed that it could work.

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About the Author

Burcu Gumuscu

Burcu Gumuscu is based BIOS Lab-on-a-Chip Group, MESA+ Institute for Nanotechnology, MIRA Institute for Biomedical Technology and Technical Medicine, University of Twente, the Netherlands.

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