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Microfluidic long DNA sample preparation from cells.

ABSTRACT: A number of outstanding problems in genomics, such as identifying structural variations and sequencing through centromeres and telomeres, stand poised to benefit tremendously from emerging long-read genomics technologies such as nanopore sequencing and genome mapping in nanochannels. However, optimal application of these new genomics technologies requires facile methods for extracting long DNA from cells. These sample preparation tools should be amenable to automation and minimize fragmentation of the long DNA molecules by shear. We present one such approach in a poly(dimethylsiloxane) device, where gel-based high molecular weight DNA extraction and continuous flow purification in a 3D cell culture-inspired geometry is followed by electrophoretic extraction of the long DNA from the miniaturized gel. Molecular combing reveals that the device produces molecules that are typically in excess of 100 kilobase pairs in size, with the longest molecule extending up to 4 megabase pairs. The microfluidic format reduces the standard day-long and labor-intensive DNA extraction process to 4 hours, making it a promising prototype platform for routine long DNA sample preparation.

SUBMITTER: Agrawal P

PROVIDER: S-EPMC6423959 | biostudies-literature | 2019 Jan

REPOSITORIES: biostudies-literature

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Microfluidic long DNA sample preparation from cells.

Agrawal Paridhi P Dorfman Kevin D KD

Lab on a chip 20190101 2

A number of outstanding problems in genomics, such as identifying structural variations and sequencing through centromeres and telomeres, stand poised to benefit tremendously from emerging long-read genomics technologies such as nanopore sequencing and genome mapping in nanochannels. However, optimal application of these new genomics technologies requires facile methods for extracting long DNA from cells. These sample preparation tools should be amenable to automation and minimize fragmentation ...[more]

PMID: 30534775

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Microfluidic long DNA sample preparation from cells.

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Microfluidic long DNA sample preparation from cells.

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