Fast genetic mapping using insertion-deletion polymorphisms in Caenorhabditis elegans

Abstract

Genetic mapping is used in forward genetics to narrow the list of candidate mutations and genes corresponding to the mutant phenotype of interest. Even with modern advances in biology such as efficient identification of candidate mutations by whole-genome sequencing, mapping remains critical in pinpointing the responsible mutation. Here we describe a simple, fast, and affordable mapping toolkit that is particularly suitable for mapping in Caenorhabditis elegans. This mapping method uses insertion-deletion polymorphisms or indels that could be easily detected instead of single nucleotide polymorphisms in commonly used Hawaiian CB4856 mapping strain. The materials and methods were optimized so that mapping could be performed using tiny amount of genetic material without growing many large populations of mutants for DNA purification. We performed mapping of previously known and unknown mutations to show strengths and weaknesses of this method and to present examples of completed mapping. For situations where Hawaiian CB4856 is unsuitable, we provide an annotated list of indels as a basis for fast and easy mapping using other wild isolates. Finally, we provide rationale for using this mapping method over other alternatives as a part of a comprehensive strategy also involving whole-genome sequencing and other methods.

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“…To our surprise, we recovered a suppressor mutant in the F1 generation of the screen and confirmed the dominant nature of this allele by subsequent backcrossing into the unmutagenized parental strain. We identified insertion-deletion polymorphisms [57] linked to the dominant suppressor mutation that mapped to the center of chromosome IV (LGIV) ( Figure 4B ). Genome-wide sequencing (GWS) of the suppressor mutant genomic DNA revealed twenty-six missense mutations within this region (Figure S4A).…”

“…Some strains were provided by the CGC, which is funded by NIH Office of Research Infrastructure Programs (P40 OD010440). Genetic mapping of the drh-1(lax257) mutation was performed as previously described [18, 19, 39, 40, 42, 77] by linkage of the gst-4p::gfp suppression of the skn-1gf(lax188) phenotype by using the InDel mapping primer set [57].…”

“…ORO-stained worms were placed on glass slides and a coverslip was placed over the sample. Worms were scored, as previously described [57, 80, 81]. Worms were scored and images were taken with LAS X software and Leica Thunder Imager flexacam C3 color camera.…”

“…Genetic mapping of the drh-1(lax257) mutation was performed as previously described [18,19,39,40,42,77] by linkage of the gst-4p::gfp suppression of the skn-1gf(lax188) phenotype by using the InDel mapping primer set [57].…”

“…Nile Red Staining. Nile Red fat staining was conducted as as previously described [57,80,81]. In brief, worms were egg prepped and allowed to hatch overnight for a synchronous L1 population.…”