Figure 3. Identification of the heterozygous point mutation E498V in exon 7 of KRT3 in the Polish MCD family. A: DNA sequencing. Electropherograms from bidirectional sequencing of KRT3 exon 7 in the proband showed a 1493 A>T (GAG>GTG) heterozygous mutation, predicting the amino aid change E498V. B: Pedigree of the studied family. The arrow indicates the proband (III-1). C: PCR-RFLP analysis. The KRT3 E498V mutation creates a recognition site for HphI. The presence of this restriction site is seen to cosegregate with MCD in this family. Upon digestion, the full sized 304 bp product is cut into bands of 260 bp and 44 bp (the latter not visible on the figure). DNA molecular weight markers are shown on the left (lane 1). The heterozygous E498V mutation (lanes 3–6) was detected in all affected family members (I-2, II-2, III-1, and III-2). The homozygous normal allele, represented by the 304 bp band (lane 2, 7, and 8), was found in unaffected family members (II-1, II-3, and III-3).