Novel method for high-throughput colony PCR screening in nanoliter-reactors.

Walser, Marcel; Pellaux, Rene; Meyer, Andreas; Bechtold, Matthias; Vanderschuren, Herve; Reinhardt, Richard; Magyar, Joseph; Panke, Sven; Held, Martin

doi:10.1093/nar/gkp160

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Novel method for high-throughput colony PCR screening in nanoliter-reactors.

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Reinhardt, Richard
High Throughput Technologies, Max Planck Institute for Molecular Genetics, Max Planck Society;

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Walser, M., Pellaux, R., Meyer, A., Bechtold, M., Vanderschuren, H., Reinhardt, R., et al. (2009). Novel method for high-throughput colony PCR screening in nanoliter-reactors. Nucleic Acids Research, 37(8), e57-e57. doi:10.1093/nar/gkp160.

Cite as: https://hdl.handle.net/11858/00-001M-0000-0010-7DED-5

Abstract

We introduce a technology for the rapid identification and sequencing of conserved DNA elements employing a novel suspension array based on nanoliter (nl)-reactors made from alginate. The reactors have a volume of 35 nl and serve as reaction compartments during monoseptic growth of microbial library clones, colony lysis, thermocycling and screening for sequence motifs via semi-quantitative fluorescence analyses. nl-Reactors were kept in suspension during all high-throughput steps which allowed performing the protocol in a highly space-effective fashion and at negligible expenses of consumables and reagents. As a first application, 11 high-quality microsatellites for polymorphism studies in cassava were isolated and sequenced out of a library of 20 000 clones in 2 days. The technology is widely scalable and we envision that throughputs for nl-reactor based screenings can be increased up to 100 000 and more samples per day thereby efficiently complementing protocols based on established deep-sequencing technologies.