CAJ | 학술논문

目的：探讨中国人尼曼-匹克病(NPD)家系发病的分子遗传学基础及SMPD1基因检测在中国人NPD家系基因诊断中的意义。方法收集3个无关的NPD家系的临床资料和血液标本，同时选取20例健康儿童的血液标本。从外周血提取基因组DNA，经聚合酶链反应(PCR)依次扩增3个家系中所有成员SMPD1基因的6个编码外显子及其侧翼内含子序列，扩增产物纯化后直接进行正反向测序，并与Genebank进行比对；将发现突变所在外显子的扩增产物通过TA克隆技术进一步证实突变的实际意义。结果家系1先证者为T107C的纯合突变，其父母均为T107C的杂合突变；家系2和家系3的所有成员均发现在SMPD1基因外显子1上有连续6个碱基缺失，即在编码序列第108碱基至第113碱基GCTGGC的缺失(c.108-113delGCTGGC)，且均为纯合突变。进一步应用TA克隆技术检测，家系2和家系3的所有成员随机挑选的单克隆测序结果均存在该突变c.108-113delGCTGGC。对20个正常对照的PCR扩增产物进行直接正反向测序，均未见以上突变。结论 SMPD1基因第1外显子上T107C的纯合突变为家系1先证者发病的分子遗传学基础，其父母是表型正常的基因突变携带者。家系2和家系3所有成员均发现在SMPD1基因第1外显子上存在c.108-113delGCTGGC的纯合突变，考虑为人类基因多态性。
목적：탐토중국인니만-필극병(NPD)가계발병적분자유전학기출급SMPD1기인검측재중국인NPD가계기인진단중적의의。방법수집3개무관적NPD가계적림상자료화혈액표본，동시선취20례건강인동적혈액표본。종외주혈제취기인조DNA，경취합매련반응(PCR)의차확증3개가계중소유성원SMPD1기인적6개편마외현자급기측익내함자서렬，확증산물순화후직접진행정반향측서，병여Genebank진행비대；장발현돌변소재외현자적확증산물통과TA극륭기술진일보증실돌변적실제의의。결과가계1선증자위T107C적순합돌변，기부모균위T107C적잡합돌변；가계2화가계3적소유성원균발현재SMPD1기인외현자1상유련속6개감기결실，즉재편마서렬제108감기지제113감기GCTGGC적결실(c.108-113delGCTGGC)，차균위순합돌변。진일보응용TA극륭기술검측，가계2화가계3적소유성원수궤도선적단극륭측서결과균존재해돌변c.108-113delGCTGGC。대20개정상대조적PCR확증산물진행직접정반향측서，균미견이상돌변。결론 SMPD1기인제1외현자상T107C적순합돌변위가계1선증자발병적분자유전학기출，기부모시표형정상적기인돌변휴대자。가계2화가계3소유성원균발현재SMPD1기인제1외현자상존재c.108-113delGCTGGC적순합돌변，고필위인류기인다태성。
Objectives To study the molecular genetics of Niemann-Pick's disease (NPD), and its implication in the diagnosis of NPD. Methods The clinical data and blood samples of three unrelated families were collected. The genomic DNA was extracted from peripheral blood. The six coding exons and their lfanking intronic sequences of SMPD1 gene in all members of three pedigrees were ampliifed by polymerase chain reaction (PCR). The SMPD1 gene sequencing results were compared with the normal sequence from Genbank to identify possible causative mutations. The ampliifcation products of exons where mutations were located were cloned into TA vector for further conifrmation. Results Family 1 proband had homozygous T107C mutation and the parents had heterozygous T107C mutation. The homozygous delete mutation (c.108-113delGCTGGC) was detected and conifrmed by TA cloning in all members of family 2 and 3. The 20 normal control members did not have this delete mutation. Conclusions The genetic basis of NPD in the proband of family 1 is the homozygous T107C mutation in SMPD1 gene, while parents in family 1 are carriers of recessive T107C mutation. The homozygous mutation c.108-113delGCTGGC exists in SMPD1 gene in all members of the family 2 and 3. This delete mutation is considered to be genetic polymorphism.