PCR-DNA直接测序检测1例单纯型大疱性表皮松解症Weber-Cockayne亚型(WC-EBS)患者角蛋白K5基因点突变

Detection of keratin 5 gene point mutation in a family with Weber Cockayne EBS by PCR and direct DNA sequencing.

单纯型大疱性表皮松解症（ＥＢＳ）是一组常染色体显性的遗传性疾病，研究表明本病存在角蛋白Ｋ５／Ｋ１４基因点突变。ＥＢＳ的各个亚型突变发生部位有一定差异，其中Ｗｅｂｅｒ－Ｃｏｃｋａｙｎｅ亚型（ＷＣ－ＥＢＳ）突变多位于Ｋ５／Ｋ１４的连接区Ｌ１－２。本研究设计了扩增Ｋ５基因Ｌ１－２区ＤＮＡ片段的引物，应用ＰＣＲ对－ＷＣ－ＥＢＳ家系的患者及未发病成员进行扩增。ＰＣＲ产物测序发现患者Ｋ５第３４６密码子发生了Ａ→Ｃ的碱基替换，导致色氨酸（ＴＡＴ）变成丝氨酸（ＴＣＴ），而未发病成员则未见有碱基突变。结果表明，通过ＰＣＲ结合ＤＮＡ直接测序不失为快速、准确检测基因突变的方法。此外，连接区在角蛋白结构中不如螺旋区重要，因而此区基因突变对角蛋白二聚体形成的影响不大，这与临床上ＷＣ－ＥＢＳ病情相对较轻是一致的。

EBS(epidermolysis bullosa simplex) is a kind of autosomal dominant inherited dermatosis, which had been proved to be related with point mutations of Keratin 5/14 genes. There were some difference of mutation spots among every subtype of EBS. For example, the mutations mainly located in Linker 1 2 domain of K5/14 in Weber Cockayne EBS(WC EBS). In this study, we amplified a 156bp DNA fragment of K5 L1 2 domain of a family with WC EBS by polymerase chain reaction. DNA sequencing showed that a A→C substitution at codon 346 of K5 gene, which resulted in tryptophan→serine. This point mutation was found only in patient, but not in unaffected family members. PCR in combination with DNA sequencing could be applied in detecting gene mutation of inherited dermatoses. In addition, we suggested that mutations in Linker domain lead minor destruction on keratin dimer's formation and cause relatively slight clinical appearance.