We observed only weak TagRFP expression in floor plate cells in some embryos after the global inversion of the gene trap cassette. We considered this expression inconclusive, and we also examined other tissues to confirm the expression of TagRFP from the inverted cassette. We detected TagRFP expression most convincingly in the ganglion cell layer (GCL) of the retina, in which robust shha mRNA expression was reported previously (Shkumatava A, et al. Development 2004), and presented the result in Figure 2—figure supplement 3.