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The spider lamb syndrome (SLS) is an autosomal recessive condition of sheep, characterized by severe bone lesions. The disease is caused by a mutation at codon 700 of the gene fibroblast growth factor receptor 3 (FGFR3), resulting in a valine (V) to glutamate (E) substitution, and leading to an abnormal bone growth. Congenital diseases as SLS may have an important impact on animal production. The objective of this work was to detect the frequency of E allele on Suffolk and H. Down breeds in three flocks from the southern region of Brazil. A total of 256 samples, 146 from two Suffolk flocks and 110 from one H. Down flocks were evaluated. The genomic DNA extracted from peripheral blood was PCR amplified and cleaved with XhoI restriction enzyme. Allele E frequency varied among the different herds: Suffolk, 6% in one flock and 3% in the other, and H. Down 6%, corresponding to 6–12% of heterozygotes for the condition. These results emphasize the need for preventive measures to minimize the birth of affected lambs. Also, the culling of animals, mainly rams, carrying the E allele will bring more efficiency to the sheep industry. These data are important also for sheep breeders, as the genotyping of the breed stock will afford more security to breeding programs and stop the spread of this undesirable allele, reducing economic losses. It is important to note that, in Brazil, the DNA test for SLS is not a routine procedure.