Extended Data Fig. 4: The HS72 enhancer can activate Shh at short-range.
From: Range extender mediates long-distance enhancer activity
(a) Transgenic E13.5 mouse embryo with the HS72 limb enhancer placed upstream of the Shh promoter and ORF and integrated at H11 safe-harbour locus (light blue). Close-up images of limbs are shown on the right. Asterisks indicate extra digits (polydactyly). Numbers of embryos with limb polydactyly in both forelimb and hindlimb buds over the total number of transgenic embryos screened are shown. FL, forelimb. HL, hindlimb. (b) Hindlimb skeletal staining of P0 ShhHS72(+2KB/wt) mice. (c) PCR gel electrophoresis confirming the correct integration of the left and right homology arms in the ShhH272(+2KB) line. One representative sample is shown for each genotype consistent with the results seen for all embryos and mice analysed in this study. (d) Model of the targeting construct used to insert the HS72 enhancer approximately 2 KB upstream of Shh. The donor vector contained two homology arms and a corresponding replaced region containing the transplanted enhancer and removing the sgRNA recognition sites (purple, 1: 5’-gggatcatgaggctggccacAGG-3’ and 2: 5’-aaaggccacatttcttcctgTGG-3’). PCR primers used for genotyping are shown as arrows. For gel source data, see Supplementary Fig. 1. See Methods for more details.
