CAJ | 학술논문

为了揭示成骨不全(Osteogenesis imperfecta, OI)Ⅰ型家系的分子遗传学发生机制，文章采用PCR-DNA直接测序法，对患儿 COL1A1和 COL1A2基因共103个外显子(E)进行突变检测。结果显示：患儿 COL1A1基因未发现任何病理性突变，而在 COL1A2基因 E19内发现一新的杂合错义突变(p.G316C)，该突变来自其父，而其母正常，其他表型正常的6位亲属也均未发现该突变；通过 DHPLC(Denaturing high performance uid chromatography)筛检，发现患儿与其父均有异常双峰，而其母和所有正常对照均为正常单峰；通过ASA(Allele specific amplification)筛检，患儿与其父均有391 bp的特异扩增带，而其母和所有正常对照均未见特异扩增带；保守性分析结果显示，该突变位点所在甘氨酸在进化上具有高度保守性；SIFT和 PolyPhen-2软件预测结果显示，新突变造成的结果是“有害的”和“很可能有害”。上述结果均说明 COL1A2基因c.946G>T/p.G316C新突变是导致OI-Ⅰ型的致病性突变，是引起患儿发病的真正内因。患儿父母若再次孕育，可在孕早期进行产前基因诊断或孕前期进行PGD(Preimplantation genetic diagnosis)予以防患。
위료게시성골불전(Osteogenesis imperfecta, OI)Ⅰ형가계적분자유전학발생궤제，문장채용PCR-DNA직접측서법，대환인 COL1A1화 COL1A2기인공103개외현자(E)진행돌변검측。결과현시：환인 COL1A1기인미발현임하병이성돌변，이재 COL1A2기인 E19내발현일신적잡합착의돌변(p.G316C)，해돌변래자기부，이기모정상，기타표형정상적6위친속야균미발현해돌변；통과 DHPLC(Denaturing high performance uid chromatography)사검，발현환인여기부균유이상쌍봉，이기모화소유정상대조균위정상단봉；통과ASA(Allele specific amplification)사검，환인여기부균유391 bp적특이확증대，이기모화소유정상대조균미견특이확증대；보수성분석결과현시，해돌변위점소재감안산재진화상구유고도보수성；SIFT화 PolyPhen-2연건예측결과현시，신돌변조성적결과시“유해적”화“흔가능유해”。상술결과균설명 COL1A2기인c.946G>T/p.G316C신돌변시도치OI-Ⅰ형적치병성돌변，시인기환인발병적진정내인。환인부모약재차잉육，가재잉조기진행산전기인진단혹잉전기진행PGD(Preimplantation genetic diagnosis)여이방환。
To uncover the molecular pathogenic mechanism of congenital osteogenesis imperfecta (OI) type I, all the 103 exons of the COL1A1 (Collagen, type Ⅰ, alpha 1) and COL1A2 (Collagen, type Ⅰ, alpha 2) genes in a child with OI type Ⅰ were screened using PCR-DNA direct sequencing. The results showed no pathological mutation in COL1A1 gene, but a novel mutation c.946G>T/p.G316C in the exon 19 of COL1A2 gene, which was inherited from her father. This mutation was not found in her mother and other six phenotypically normal relatives. By denaturing high perfor-mance liquid chromatography (DHPLC) screening, the abnormal double-peak was visualized in PCR products of exon 19 of COL1A2 gene in the proband and her father, while the normal single-peak was shown in those of her mother and all the healthy controls. Using allele specific amplification (ASA) screening, a specific band of 391 bp in COL1A2 exon 19 was amplified only in the proband and her father, but not in other samples. The amino acid encoded by the mutation site is evolutionarily highly conserved, and this mutation was a“damaging”or“probably damaging”factor to OI type Ⅰ, based on the predicting results using SIFT and Polyphen-2 softwares. In conclusion, the novel c.946G>T/p.G316C mutation in COL1A2 gene is a pathogenic mutation that could result in OI type Ⅰ. If the couple wants to get pregnant again, it is necessary to screen the mutation site in COL1A2 gene through the prenatal genetic diagnosis in the first trimester or through preimplantation genetic diagnosis (PGD) in the progestation.