Lissens W, De Meirleir L, Seneca S, Benelli C, Marsac C, Poll-The B T, Briones P, Ruitenbeek W, van Diggelen O, Chaigne D, Ramaekers V, Liebaers I
Department of Medical Genetics, University Hospital, Brussels, Belgium.
Hum Mutat. 1996;7(1):46-51. doi: 10.1002/(SICI)1098-1004(1996)7:1<46::AID-HUMU6>3.0.CO;2-N.
Most of the mutations causing deficiency of the pyruvate dehydrogenase (PDH) complex are in the X-linked E1 alpha gene. We have developed a rapid screening method for the detection of mutations in this gene using reverse transcription of total RNA, polymerase chain reaction amplification of the whole coding region of the gene and single-strand conformation polymorphism (SSCP) analysis. With this method, we studied eight patients with a PDH complex deficiency, using cultured fibroblasts. In all patients, aberrant SSCP patterns were found and, after sequencing of the corresponding fragments, we were able to identify six new mutations and two mutations already described previously. The mutations are point mutations leading to amino acid substitutions (5) and direct repeat insertions (3). The presence of the mutations was confirmed in genomic fibroblast DNA. The 4 female patients were shown to carry both a normal and a mutated E1 alpha gene.
大多数导致丙酮酸脱氢酶（PDH）复合体缺乏的突变存在于X连锁的E1α基因中。我们开发了一种快速筛选方法，通过对总RNA进行逆转录、对该基因的整个编码区进行聚合酶链反应扩增以及单链构象多态性（SSCP）分析来检测该基因中的突变。利用这种方法，我们使用培养的成纤维细胞研究了8例PDH复合体缺乏的患者。在所有患者中均发现了异常的SSCP模式，对相应片段进行测序后，我们能够鉴定出6个新突变和2个先前已描述的突变。这些突变是导致氨基酸替代的点突变（5个）和直接重复插入（3个）。在基因组成纤维细胞DNA中证实了这些突变的存在。4名女性患者被证明同时携带正常和突变的E1α基因。
