Figure 2

Inducible Cre constructs do not label limb muscle. (a) Representation of the Car-ERCreER/CALNL-GFP construct, where Car promoter is driving the expression of the tamoxifen-inducible Cre recombinase. On the same vector but in opposite direction, we placed a reporter construct composed of the constitutive CAG promoter driving the expression of a floxed neo gene. Downstream of it, there is the eGFP gene that is only expressed when Cre is able to remove the floxed neo gene. (b, c) After two weeks of daily intraperitoneal tamoxifen injections, only few myofibres were generally labelled in the tails (b) and less than 1% of transgenics had any label in the hindlimb (c) of Car-ERCreER/CALNL-GFP F₁ and F₂ transgenics (arrows: faint GFP⁺ myofibres). n > 100 animals. (d) Representation of the Car-nCre, Hsp-nlcCre and CALNL-GFP constructs. Here, the Cre recombinase is split in two inactive fragments (nCre and nlcCre). In this system, Cre is only active when both fragments are co-expressed. To make the system inducible and muscle-specific, we cloned the nCre fragment under the Car promoter and the nlcCre fragment under a heat-shock-inducible promoter (Hsp70). (e-g) After heat-shock treatments of F₁ tadpoles, we generally observed strong expression of GFP in the trunk (arrow) but no expression in the hindlimbs (compare e with e'. Arrowheads point to the unlabelled hindlimb). We also observed that the number of labelled myofibres was lower in the middle of the tail (g) and even lower closer to the tip of the tail (f). n > 100 animals.