Fig 1.
Pigmentation and banding patterns display marked variation in C. nemoralis.
A. Representative shell colour morphs of C. nemoralis. Shell background colour can range from white to yellow and pink to dark orange or brown. Banding patterns include zero to five bands that can merge or be distinct. B. A shell with a yellow background and a typical banding pattern. The lateral pigmented band at the leading edge of the shell that indicates a fully differentiated snail is indicated with a horizontal red arrow. C. Banding patterns in the shell are reflected in the pigmentation of the underlying mantle tissue, and facilitate the isolation of populations of cells for separate RNA extractions and gene expression profiling. The mantle tissue presented here was responsible for both constructing and pigmenting the shell shown in B. The individual pigmented bands in the mantle are numbered and correspond to those indicated on the shell.
Table 1.
Primer sequences and amplicon sizes for all used reference genes.
Fig 2.
A schematic representation of our sampling and experimental design and data analysis.
RNA was extracted from four different tissues derived from six snail replicates across two seasons. The four independent tissues (and five combinations thereof) and three algorithms were used to rank gene performance, and these results were then combined to find the overall best ranked genes.
Fig 3.
Expression stability ranking across all tissues within each individual snail.
Means of ranked data generated by three algorithms (BestKeeper, geNorm and NormFinder). Error bars indicate standard error.
Fig 4.
Expression stability ranking within each tissue type.
Means of ranked data generated by three algorithms (BestKeeper, geNorm and NormFinder). Error bars indicate standard error.
Fig 5.
Expression stability ranking of combined tissue types.
Body (Foot and Head); Mantle (Mantle pigmented and unpigmented); M+F (Mantle and Foot); M+H (Mantle and Head); All (all tissue sets). Means of ranked data generated by three algorithms (BestKeeper, geNorm and NormFinder). Error bars indicate standard error.