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11.
  • Schnorrer, Frank, et al. (författare)
  • Positional cloning by fast-track SNP-mapping in Drosophila melanogaster
  • 2008
  • Ingår i: Nature Protocols. - 1754-2189 .- 1750-2799. ; 3:11, s. 1751-1765
  • Tidskriftsartikel (refereegranskat)abstract
    • Positional cloning of chemically induced mutations is the rate-limiting step in forward genetic screens in Drosophila. Single-nucleotide polymorphisms (SNPs) are useful markers to locate a mutated region in the genome. Here, we provide a protocol for high-throughput, high-resolution SNP mapping that enables rapid and cost-effective positional cloning in Drosophila. In stage 1 of the protocol, we use highly multiplexed tag-array mini-sequencing assays to map mutations to an interval of 1-2 Mb. In these assays, SNPs are genotyped by primer extension using fluorescently labeled dideoxy-nucleotides. Fluorescent primers are captured and detected on a microarray. In stage 2, we selectively isolate recombinants within the identified 1-2 Mb interval for fine mapping of mutations to about 50 kb. We have previously demonstrated the applicability of this protocol by mapping 14 muscle morphogenesis mutants within 4 months, which represents a significant acceleration compared with other commonly used mapping strategies that may take years.
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