Mutations in the GJB2 gene encoding the gap‐junction protein connexin 26 have been identified in many patients with childhood hearing impairment (HI). One single mutation, 35delG (30delG), accounts for up to 70% of all analyzed European patients with autosomal recessive inherited HI and 10% of patients with HI of unknown origin, respectively. We screened 188 control individuals and 342 German patients with non‐syndromic sporadic HI for the 35delG, compound heterozygosity and other GJB2 mutations by PCR, restriction enzyme based screening, SSCP and sequencing. In all patients, non‐progressive hearing impairment varied from moderate to profound involving all frequencies. This study revealed one novel silent mutation (438C/T), three novel gene variants resulting in amino acid substitutions (K112E, T123S, K223R) and two novel HI‐related mutations (I82M, 313del14). © 2002 Wiley‐Liss, Inc.