基于目的基因捕获的高通量测序技术在遗传代谢病诊断中的应用  

Application of high throughput targeted exome sequencing in the molecular diagnosis of inherited metabolic diseases

作　　者：郝虎[1] 石聪聪[2] 吴鹰军[3] 王弘 李思涛[1] 蔡尧[1] 董玛拉 马艳梅[2] 肖昕[1] 

机构地区：[1]中山大学附属第六医院新生儿科,广州510655 [2]中山大学附属第六医院遗传代谢病实验室,广州510655 [3]广州医科大学附属第四医院检验科 [4]广州市天河区妇幼保健院儿科

出　　处：《中华实用儿科临床杂志》2015年第22期1721-1724,共4页Chinese Journal of Applied Clinical Pediatrics

基　　金：广东省科技计划项目（2014A020212133）;广东省天河区科技计划项目（2013KW038）

摘　　要：目的探讨基于目的基因捕获的高通量测序技术对串联质谱联合气相色谱-质谱技术在遗传代谢病（IMD）诊断中的应用价值。方法收集中山大学附属第六医院遗传代谢病实验室通过串联质谱联合气相色谱-质谱技术检测的疑似阳性10例患儿及其父母外周血样本，通过包含有153个常见的IMD致病基因捕获探针的高通量测序技术对患儿进行基因检测，采用Sanger测序技术对患儿阳性突变位点进行验证，并对其父母相应的位点进行检测。结果10例疑似IMD患儿，通过高通量测序技术确诊8例，包括全羧化酶合成酶缺陷症3例，甲基丙二酸血症mut型、甲基丙二酸血症cblB型、枫糖尿症Ⅰa型、鸟氨酸氨甲酰转移酶缺乏症、Citrin缺陷症各1例。其中有1例经串联质谱联合气相色谱-质谱诊断为异戊酸血症的患儿，其基因检测发现1个错义突变（c．158G〉C，p．R53P）和1个同义突变（c．732C〉T，p．D244D）；另外1例疑诊为Citrin缺陷症的患儿，其基因检测发现1个错义突变（c．1156G〉A，p．G386S）和1个同义突变（c．1194A〉G，p．L39SL），这2例基因检测结果在理论上尚不能确诊。10例患儿中鸟氨酸氨甲酰转移酶缺乏症患儿的致病突变属于自身新发生的基因突变，而其他患儿的致病突变均遗传于各自的父母。此外，高通量测序所检测的结果均经Sanger测序验证，二者结果全部一致。结论基于目的基因捕获的高通量测序技术在疑似IMD的检测中结果精确，不仅可为临床医师提供可靠的分子诊断依据，还能精准地进行疾病分型，有效地为后续临床治疗和遗传咨询提供依据。Objective To evaluate the application value of high throughput targeted exome sequencing in inherited metabolic disorders（IMD） diagnosed by tandem mass spectrometry combined with gas chromatography -mass spectrometry. Methods Ten cases of peripheral blood were collected from the parents and children with suspected inborn errors of metabolism detected by tandem mass spectrometry combined with gas chromatography mass spectrometry in the testing center of the Sixth Affiliated Hospital of Sun Yat - Sen University. One hundred and fifty - three genes related to genetics metabolic disorders were analyzed to obtain the positive mutations utlizing the high throughput targeted exome sequencing technology. Then all the mutations were validated by Sanger sequencing, as well as their parents＇ corresponding sites. Results Eight of the 10 patients with suspected IMD were confirmed by high throughput sequencing targeted exome sequencing. Among the 8 patients, 1 patient with methylmalonic academia - mut type, 1 patient with methylmalonic academia- cblB type, 1 patient with maple syrup urine disease - Ⅰ a type, 1 patient with ornithine carbamoyhransferase deficiency, 1 patient with citrin deficiency, and 3 patients with holocarboxylase synthetase deficiency. However,there were still 2 cases which could not be verified, 1 patient with the suspected isovaleric acidaemia diagnosed by gas chromatography - mass spectrome spectrometry who had a missense mutation c. 158G 〉 C （p. R53P） and a same sense mutation c. 732C 〉 T（ p. D244D） ,the other one with citrin suspected deficiency had a missense mutation c. 1156G 〉 A（ p. G386S） and a same sense mutation c. 1194A 〉 G（ p. L398L）. Moreover,the positive gene sites of the patient with ornithine carbamoyltransferase deficiency （ X - linked recessive inheritance） mutated spontaneously which differed from the other 9 cases inherited from their own parents. Besides, all the data of high - throughput sequencing were confirmed by Sanger sequencing and in accordan

关 键 词：高通量测序 遗传代谢病 串联质谱 气相色谱一质谱 

分 类 号：R440[医药卫生—诊断学] R722.1[医药卫生—临床医学]