(A) The snail genera used in this study (image credits: Lymnaea [E. de Roij], Biomphalaria and Physa [creative commons], Partula and Euhadra [A.D.]).

(B) 3,403 offspring were used to infer the recombination breakpoints that bound the D locus. Numbers of mapped recombinants for 1,507 sinistral (dd) snails are shown on the right and for 1,896 dextral (DD or Dd) on the left. The sinistral mutation must be between loci b6 and b12 (shaded), a region that spans 267 kb (not to scale).

(C) Boxplots show normalized relative quantities (NRQs), on log scale, of quantitative real-time PCR assays of transcripts of three candidate genes and one control (Larp2/3 1a) in single-cell egg samples from dextral homozygote (DD), dextral heterozygote (Dd), and sinistral recessive homozygote (dd) individuals. Significant differences in expression were detected for Ldia2 only (DD:dd, p = 0.002; DD:Dd and Dd:dd, p = 0.004).

(D) WMISH of maternal Ldia transcripts in early, dextral L. stagnalis embryos.

(E) Schematic showing two hypotheses for the evolution of chirality in three snail families (dextral = blue; sinistral = red). Either sinistrality evolved once from a dextral ancestor, with the ancestral Lymnaeid reverting to dextral (bottom), or sinistrality evolved twice (top).