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

Nucleic Acids Res.. 2009-05; 
Walser Marcel, Pellaux Rene, Meyer Andreas, Bechtold Matthias, Vanderschuren Herve, Reinhardt Richard, Magyar Joseph, Panke Sven, and Held Martin. ETH Zurich, Institute of Process Engineering, BioProcess Laboratory (BPL), Zurich, Switzerland.
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摘要

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... More

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