Marc Hammarlund et al. (2002) West Coast Worm Meeting "Mapping a new class of Ric mutants using an improved SNP mapping technique"

Tracey Harrach, Erik M Jorgensen, Marc Hammarlund

[West Coast Worm Meeting, 2002]

In an effort to identify additional molecules that function in synaptic transmission we have conducted several large screens for Aldicarb resistance. Aldicarb is an inhibitor of acetylcholinesterase and mutants that secrete less neurotransmitter are resistant to this drug. The mutants obtained from these screens fall into two classes: those that are uncoordinated, and those that are not uncoordinated.

M Wayne Davis et al. (2005) International Worm Meeting "High-throughput SNP mapping primers and techniques"

Erik M Jorgensen, Tracey Harrach, Marc Hammarlund, Patrick Hullett, M Wayne Davis, Shawn Olsen

[International Worm Meeting, 2005]

Single nucleotide polymorphism (SNP) mapping against CB4856 has quickly become an indispensable technique in the worm field since its introduction by Stephen Wicks et al. in 2001 (Nat Genet 2001 28: 160-164). Because SNPs are present every few thousand base pairs, SNP mapping allows mapping to extremely narrow intervals. Because mapping is done in an essentially wild-type background, very subtle or complex phenotypes can be mapped that could not be mapped using visible mutations. Typically, SNPs that alter a restriction enzyme recognition site (snip-SNPs) have been detected by running digested PCR products on agarose gels. This technique is reliable and readily accessible to any molecular biology lab; however, it requires substantial hands-on time for each reaction, as each SNP typically requires different PCR cycling and restriction digestion conditions. A number of methods have been published that provide automated analysis of SNP genotypes, including florescence polarization (Swan et al. 2002 Genome Res 12: 1100-1105) or capillary electrophoresis (Zipperlen et al. 2005 Genome Biol 6: R19). Although these methods have the advantage of reducing hands-on time for determining SNP genotypes, they require investment in expensive equipment that is not common in many molecular biology labs. In order to provide a more efficient detection protocol for snip-SNPs, we have developed a set of 48 primer pairs that detect SNPs evenly spaced across the C. elegans genome, and that work under identical PCR and restriction digestion conditions. We present a scheme using these reagents to quickly and reliably map mutations using high-throughput equipment and techniques. We have used our SNP mapping techniques to map many phenotypically subtle or genetically complex mutants, including a subtle behavioral defect resulting from three independent mutations. In fact our techniques are simple and relatively inexpensive enough to have been used successfully by undergraduates in a teaching laboratory setting.