Ohshima T, Takahashi J, Tohgi H, Sakuragawa N
Division of Inherited Metabolic Disease, Iwate Medical School.
Rinsho Shinkeigaku. 1994 Jan;34(1):1-4.
We performed the genomic analysis of arylsulfatase A (ASA) gene in five Japanese patients with adult-type metachromatic leukodystrophy (MLD) including two sibling cases. Sequencing of amino acid coding region of ASA gene of proband case of family A disclosed 426Pro (CCG)-->Leu (CTG) mutation, which was reported to be frequently found in Caucasian patients with late-onset MLD. We developed mismatch primer PCR/RFLP method for detection of this mutation. If 426Pro-->Leu mutation exists in genomic DNA, Pst I site is newly created by PCR with a 3'-primer mismatched at one nucleotide. Genomic analysis of family A members using this method revealed that younger patient was homozygote of 426Pro-->Leu mutation and patient's parents and her younger brother were heterozygotes, which were confirmed by sequencing of exon 8 of ASA gene. Screening of this mutation using mismatch primer PCR/RFLP method was performed in one sibling case and one autopsy case. This point mutation was found in the sibling case. These results showed the possibility of world-wide spread of 426Pro-->Leu mutation in late-onset MLD patients and usefulness of our mismatch primer PCR/RFLP method for screening of this mutation.
我们对5名成年型异染性脑白质营养不良（MLD）的日本患者（包括2例同胞患者）进行了芳基硫酸酯酶A（ASA）基因的基因组分析。对A家族先证者的ASA基因氨基酸编码区进行测序，发现了426Pro（CCG）→Leu（CTG）突变，据报道该突变在晚发型MLD的白种人患者中经常出现。我们开发了错配引物PCR/RFLP方法来检测这种突变。如果基因组DNA中存在426Pro→Leu突变，那么在使用一个核苷酸错配的3'引物进行PCR时会新产生Pst I位点。使用该方法对A家族成员进行基因组分析发现，年轻患者是426Pro→Leu突变的纯合子，患者的父母和弟弟是杂合子，这通过ASA基因第8外显子的测序得到了证实。在1例同胞病例和1例尸检病例中使用错配引物PCR/RFLP方法对该突变进行了筛查。在同胞病例中发现了这种点突变。这些结果表明426Pro→Leu突变在晚发型MLD患者中可能在全球范围内传播，并且我们的错配引物PCR/RFLP方法在筛查这种突变方面具有实用性。
