Objective To identify a genetic disease of pretibial epidermolysis bullosa (PEB).Method Genomic DNA extracted from peripheral blood from the patients and other unaffected family members by the phenol-chloroform extraction method.The mutations of the COL7A1 gene was undertaken by sequencing.Sanger sequencing using customized primers was carried out to determine the candidate causative mutation in all available clinically affected subjects as well as to screen unaffected members in the family for co-segregation analyses.The mutation analysis in the gene was further evaluated in 200 ethnically matched normal controls by denaturing high-performance liquid chromatography (DHPLC).Results A novel heterozygous nucleotide G>T transition at position 6101 in exon 73 of COL7A1 was detected, which resulted in a glycine to valine substitution (G2034V) in the triple-helical domain of type-Ⅶ collagen.Conclusion This is the first report to show that one mutation caused a broad range of severity of disease in one family with PEB.These data suggest that c.6101G>T may influence the phenotype of PEB.They also contribute to the expanding database on COL7A1 mutations.