-------- Original Message -------- Subject: Morphometrics of small, variable specimens--embryos Date: Tue, 29 Mar 2011 11:15:26 -0400 From: [email protected] To: [email protected] Hello all, I have a second question to ask. Again, I am doing some analyses of midgestational mouse embryos. The different groups are nearly genetically identical. I am looking for morphometric differences due to differences in genotype at a single or at two loci. However, the effect of genotype will be mixed up among the variation in the data owing to heterogeneity of developmental progress among the specimens. The level of ontogenetic variation is large. I generally take the procrustes coordinates (the symmetric component when I can) and regress on centroid size. I also regress those residuals on developmental stage by counting somites. This last set of residual coordinates I use as the data to generate the covariance matrix. But I still notice that the specimens tend to non randomly clump by litter in scatter plots (like PCA). I think of this as a "litter effect," but I imagine that this effect owes to significant residual ontogenetic variation in the data. For example, when I regress PC1 against developmental stage, an orderly curve results. The most and least developmentally advanced specimens occupy the same end of the PC, with the intermediates scattered in between. Similarly, when I regress the Procrustes residuals against somite number, a "growth curve" results. It has the same form as a growth curve, being horizontal at the low and high ends and positively sloped in between. The intermediately staged embryos dominate the linear trend. This makes sense, since the dominate trend of PC1 is the difference between the intermediate embryos and the rest of the sample, the least and most developmentally advanced. The shape changes at the extremes are "not seen clearly" be the analysis. They are collapsed together. I think this means that the allometry of embryonic growth is, while a large component, not the only significantly dominating component of the ontogenetic variation in the sample. The non-linear component of shape variation is hard to get rid of in embryos. These patterns persist even when different developmental time spans are considered. Breaking up a sequence into smaller sequences does not seem to be equivalent to isolating those specimens along the linear trend from those that are not. The same non-linear "growth curve" results after regressing procrustes residuals against somite number for the sub sample of intermediates alone, though it may not be as pronounced. So my question is, given that my different groups are nearly identical genetically, I can expect that they will be very similar looking in the analysis. If I cannot see a separation of the genotypes along PCs or along a discriminant function, I feel like I can argue that no morphometric difference has been detected, despite the presence of residual ontogenetic variation, which would serve to actually separate my groups, since my samples are never exactly the same in developmental stage composition. Would you agree that the data are biased toward finding a shape difference between the two groups, biased because of remaining shape variation due not to genotype but to developmental heterogeneity? So that a negative result of no shape difference detected should be interpreted as particularly robust? Thanks. Eric [email protected] University of Calgary Faculty of Medicine
