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Positional cloning by fast-track SNP-mapping in Drosophila melanogaster
Frank Schnorrer1, Annika Ahlford, Doris Chen
1Research Institute of Molecular Pathology, Dr Bohr-Gasse 7, A-1030 Vienna, Austria. schnorrer@biochem.mpg.de
Nature Protocols
|October 25, 2008
Summary
This study introduces a rapid, cost-effective protocol for positional cloning in Drosophila using high-resolution single-nucleotide polymorphism (SNP) mapping. The method significantly accelerates the identification of chemically induced mutations, speeding up genetic screens.
Area of Science:
- Genetics
- Molecular Biology
- Developmental Biology
Background:
- Positional cloning of mutations is a bottleneck in forward genetic screens.
- Single-nucleotide polymorphisms (SNPs) are crucial for genomic region identification.
- Efficient mutation mapping is vital for Drosophila research.
Purpose of the Study:
- To develop a high-throughput, high-resolution SNP mapping protocol.
- To enable rapid and cost-effective positional cloning in Drosophila.
- To accelerate the process of identifying chemically induced mutations.
Main Methods:
- Utilizing highly multiplexed tag-array mini-sequencing assays for initial mapping (1-2 Mb intervals).
- Employing primer extension with fluorescently labeled dideoxy-nucleotides for SNP genotyping.
- Implementing selective isolation of recombinants for fine mapping (to ~50 kb).
Main Results:
- The protocol enables mutation mapping to 1-2 Mb intervals in stage 1.
- Fine mapping to approximately 50 kb is achieved in stage 2.
- Successfully mapped 14 muscle morphogenesis mutants within 4 months.
Conclusions:
- The developed SNP mapping protocol significantly accelerates positional cloning in Drosophila.
- This method offers a substantial improvement over traditional mapping strategies.
- The protocol enhances the efficiency of forward genetic screens.
