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Targeted mutagenesis in a human-parasitic nematode

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CRISPR-Cas9 targeting of the Ss-unc-22 gene results in iL3s with an uncoordinated phenotype.

(A-B) Time-lapse images of wild-type iL3s (A) vs. unc F1 iL3s (B) swimming in a water droplet. Wild-type iL3s showed continuous rapid movement in water; unc F1 iL3s experienced intermittent bouts of twitching, paralysis, and uncoordinated movement. For A and B, red lines indicate iL3 trajectories. Scale bars = 200 μm. (C) Swimming distance for wild-type iL3s vs. unc F1 iL3s over a 10-s period. unc F1 iL3s swam shorter distances relative to wild-type iL3s. ***P<0.001, Mann-Whitney test. n = 21–23 trials for each population. (D) Average crawling speed for wild-type iL3s vs. unc F1 iL3s over a 20-s period. unc F1 iL3s showed reduced crawling speeds relative to wild-type iL3s. **P<0.01, unpaired t test with Welch’s correction. n = 30–32 trials for each population. (E) Maximum crawling displacement for wild-type iL3s vs. unc F1 iL3s over a 5-min period. unc F1 iL3s traversed less distance than wild-type iL3s. **P<0.01, Mann-Whitney test. n = 26 trials for each population. For C-E, graphs show medians and interquartile ranges. unc F1 iL3 data for B-E were obtained from plasmid vector delivery of CRISPR-Cas9 constructs at Ss-unc-22 site #1.

Fig 2

doi: https://doi.org/10.1371/journal.ppat.1006675.g002