摘要
目的探讨分子诊断技术在新生儿葡萄糖－6－磷酸脱氢酶（G6PD）缺乏症诊断中的应用，鉴定G6PD基因新的突变位点。
方法 收集重庆医科大学附属儿童医院新生儿疾病筛查中心2013年12月至2014年6月新生儿G6PD缺乏症筛查阳性的患儿231例，采用PCR和Sanger测序法对新生儿G6PD缺乏症筛查阳性的患儿进行G6PD基因突变检测，以期明确本地区的突变热点；通过对筛查阳性标本G6PD活性比值法检测结果与分子诊断结果的比较，分析酶学表型与基因型的关系，结合突变的功能预测，鉴定G6PD基因新的突变。
结果 本研究联合比值法和分子诊断在筛查阳性的患者中共确诊G6PD患儿125例，其中比值法确诊102例，阳性率81.6%，分子诊断确诊123例，突变检出率98.4%。比值法可疑阳性的16例患儿中，分子诊断检出突变11例。113例比值法阴性的标本中检测出突变12例，突变检出率10.6%。基因测序发现2个新突变位点，c.29C〉A和c.361C〉T，其中c.29C〉A的检出率高达7.3% （9/123），可能成为中国人群G6PD基因一个新的突变热点。
结论 新生儿G6PD缺乏症活性比值法检测容易漏诊女性杂合子；G6PD基因存在新的突变，且突变热点存在地域差异；基于DNA测序分析的分子诊断是进行新生儿G6PD缺乏症确诊最有价值的方法。
ObjectiveTo evaluate the clinical significance of molecular diagnosis for detecting neonatal glucose-6-phosphate dehydrogenase （G6PD） deficiency and to identify the novel mutations in G6PD gene.
MethodsTotally 231 newborn blood samples with decreased G6PD deficiency assessed by first-line newborn screening were recruited from Children＇s Hospital, Chongqing Medical University. G6PD gene mutation analysis was performed by PCR following by Sanger DNA sequencing, and then hotspot mutation sites were confirmed. The relationship between the enzymatic phenotype and genotype was evaluated by comparing the results between the G6PD activity detection and the molecular diagnosis. Novel variations were mutation predicted to identity the pathogenic mutations.
ResultsTotally 125 G6PD patients were identified in present study group, among which 102 patients were classified as positive by G6PD/6PGD quantitative tests, giving out a detecting rate of 81.6%. The 123 patients were confirmed by molecular diagnosis with a detecting of 98.4%. Within the 16 probably positive cases, 11 definite mutations were confirmed by molecular diagnosis. DNA sequencing also confirmed another group of 12 patients from the 113 negative samples, giving a detecting rate of 10.6%. Two novel mutations of G6PD gene, c. 29C〉A and c. 361C〉T were found in the study, of which, c. 29C〉A was observed in 7.3% （9/123） of the samples and might be a new hotspot site in Chinese population.
ConclusionsQuantitative ratio testing may be not efficient enough for G6PD deficiency diagnosis in patients carrying heterozygous mutations. Novel mutations can be found in G6PD gene and obviously there are regional diversities in the frequency of hotspot mutations for G6PD deficiency. Therefore, DNA sequencing based molecular testing may serve as the most valuable method for the diagnosis of neonatal G6PD deficiency.
出处
《中华检验医学杂志》
CAS
CSCD
北大核心
2016年第11期843-847,共5页
Chinese Journal of Laboratory Medicine
基金
基金项目：国家自然科学基金（21305165,81373444）
重庆市卫生局科学基金（2012-1-049,2013-1-028）
重庆市科委项目（Cstc2015shmszx120012）
关键词
葡糖磷酸脱氢酶缺乏
葡糖磷酸脱氢酶
序列分析
DNA
分子诊断技术
突变
新生儿筛查
Glucosephosphate dehydrogenase deficiency
Glucosephosphate dehydrogenase
Sequence analysis, DNA
Molecular diagnostic techniques
Mutation
Neonatal screening
