Description

Genetic balancer strains are frequently used to maintain populations of C. elegans mutants that would otherwise die as homozygotes. Additionally, fluorescently-marked strains can be used to identify cross-progeny for use in assays where heterozygotes are of particular interest. In our case, we wanted to utilize a fluorescently-marked but otherwise wildtype strain to obtain marked cross-progeny from a mating. Our mutant of interest displayed altered pharyngeal pumping, so we wanted to assess heterozygotes in the same assay and establish the recessive nature of the phenotype. We happened to have the TY4236 strain growing in the lab, which not only expresses bright pharyngeal and gut GFP from the mIs10 insertion but also contains him-8(e1489), making the experiment quite simple. Due to the bright GFP signal in the pharynx we decided to assess the pharyngeal pumping of the strain itself before using it as a marker in an experiment that heavily implicates the pharynx.

To our surprise, this otherwise-healthy strain displayed highly variable pharyngeal pump frequency, decreased average pump duration, and increased average inter-pump interval (IPI) when compared to N2 (Fig 1A-C). This prompted us to test multiple other options, including some that also express GFP in the pharynx (tmC5::Venus, hT2::gfp, nT1::gfp) as well as two that don’t [myo-3p::gfp::GFP(mit), tdTomato::H2B]. Two balancers with bright pharyngeal GFP, hT2::gfp and nT1::gfp, displayed moderate pumping phenotypes even as heterozygotes (Fig 1A-C). Interestingly, the pharyngeal pumping phenotypes do not correlate exactly with fluorescence localization; tmC5::Venus was completely unaffected and myo-3p::gfp(mit) myo-3::GFP(mito) had slightly but significantly reduced mean pump duration (Fig 1B). Besides N2, each strain was only assayed once and therefore these results should be treated as preliminary. Additionally, since these strains were all created in different labs across the world and in some cases across the span of many years, it is possible that the pumping phenotype is due, in part, to the genetic background of the strain and not entirely due to the insertion of a fluorescent tag.

Pharyngeal pumping phenotypes can indicate upstream nervous system or muscular alterations and can lead to downstream outcomes like altered metabolism and lifespan (Raizen et al., 1995). Regardless of the experimenter’s interest in characterizing pharyngeal pumping, these fluorescent strains should be assessed for possible subtle phenotypes that could potentially confound the main phenotype of interest.