The advent of high coverage and low cost sequencing technologies has allowed for newer and more powerful approaches in molecular and population genetics. Transposon sequencing, where genome-saturated mutant populations’ allele frequencies are measured before and after selection, functionally characterizes each and every gene in the genome in a single experiment. The approach has been successfully applied to a variety of phenotypes in a variety of unicellular systems: growth and motility in E. coli, synthetic genetic interactions in yeast, and pathogen-resistance in mammalian cell lines. However, transposon insertion typically produces null alleles, which can be valuable to identify gene function, but evolutionary insight relies on identification of naturally occurring polymorphisms affecting the trait of interest. Genome-wide association studies (GWAS) can be used to study the effect of natural genetic variation on a trait, but they grow prohibitively expensive if the number of individuals to genotype and phenotype becomes large. Techniques such as pool-GWAS, where individuals are pooled prior to genotyping, and restriction site-associated DNA sequencing (RAD-seq), where a subset of the genome is targeted for sequencing, have lowered the cost and increased the power of GWAS.
Here we describe the application of transposon sequencing and pool-GWAS (with RAD-seq genotyped natural variants) in the whole metazoan model, C. elegans. Transposon sequencing has not been previously implemented in an animal model. Specific challenges we have solved include the lack of high-throughput transformation (in contrast to unicellular models) and creation of a representative transposon-insertion sequencing library. We are using our new transposon-sequencing and pool-GWAS methods to study the genetic underpinnings of starvation survival in the nematode. When a C. elegans larva hatches in the absence of food its development must be halted and its metabolism changed, requiring precise coordination within cells and between tissues. We present preliminary data of a transposon sequencing screen for starvation survival. Furthermore, we present preliminary data of a pool-GWAS experiment of 96 C. elegans wild isolates for starvation survival.