Fig. 3

sde1 encodes a laminin beta1 paralog.

(A) Analysis of genomic homozygosity in sde1 mutants. (B) Log likelihood analysis using SNPtrack. (C) High-resolution mapping using linked SNP markers. Three and five recombinants were found for SNP32559436 and SNP32681121, respectively, on each side of a 121 kb region. After genotyping 453 adult animals, no recombinants were identified for a novel SNP at position 32605161. (D) Sanger sequencing readouts from wild-type and sde1 cDNA. SNP32605161 is within the coding region of the gene laminin beta 1a (lamb1a), causing a leucine to proline change. (E) Amino acid alignment across distant species. Red star marks the location of the leucine. Differential gray scale indicates level of conservation across listed species. (F) Cartoon depicting major structural domains in Lamb1a. Blue and red arrows indicate the locations of the sde1 mutation, along with two previously identified alleles gup^m189 and s804. Lam NT, Laminin N-terminal domain; Blue hexagons, Laminin-type epidermal growth factor-like domain; CC, uncharacterized coiled-coil domain. (G) sde1 embryos incubated at 31°C have shortened trunks. Representative embryos from an sde1 x sde1/+ cross. Embryos were transferred to 31°C at 3 hours post-fertilization (hpf). Images were acquired at 28 hpf. Approximately 48% of embryos (32 out of 67) showed phenotypes representative of the gup^m189 mutation after the temperature shift, consistent with expected Mendelian ratio. The phenotype and the ratio were consistent across three independent crosses. (H) Complementation tests showing both gup^m189 and s804 alleles fail to complement the 7 dpa regeneration defects of the sde1 mutation in adult animals, yielding expected ratios (~50%; n = 65 and 30). Red dashed lines indicate plane of amputation. Scale bars, 1 mm.