上海交通大学学报（医学版）

上海交通大学学报（医学版）

• 论著（基础研究） • 上一篇    下一篇

法洛四联症患儿NOTCH2基因3′UTR区域突变分析

冯超1，唐宁1，方绍海1，徐月娟1，李奋2，徐让1   

  1. 上海交通大学 医学院 1.附属新华医院科研中心， 上海 200092; 2.附属上海儿童医学中心， 上海 200127
  • 出版日期:2015-04-28 发布日期:2015-04-29
  • 通讯作者: 徐让, 电子信箱: rang_xu@hotmail.com。
  • 作者简介:冯超（1984—）， 女， 助理研究员， 博士; 电子信箱: workorderfc@126.com。
  • 基金资助:

    国家自然科学基金(81070135)；国家重点基础研究发展计划(“973”计划)(2010CB529501)

Analysis of mutation in 3′UTR region of NOTCH2 gene of children with tetralogy of Fallot disease

FENG Chao1, TANG Ning1, FANG Shao-hai1, XU Yue-juan1, LI Fen2, XU Rang1   

  1. 1.Scientific Research Center, Xinhua Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200092, China; 2.Shanghai Children's Medical Center, Shanghai Jiao Tong University School of Medicine, Shanghai 200127, China
  • Online:2015-04-28 Published:2015-04-29
  • Supported by:

    National Natural Science Foundation of China, 81070135; National Program on Key Basic Research Project of China, “973”Program, 2010CB529501

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摘要：

目的 筛查法洛四联症（TOF）患儿中NOTCH2基因3′UTR区域存在的基因突变，探讨该区域突变影响NOTCH2基因表达的可能机制。方法 选取152例诊断明确的TOF患儿（TOF组）为研究对象，107名健康儿童为对照组(CON组)，PCR扩增NOTCH2基因3′UTR区域序列，所有扩增片段均进行双向测序。将所测NOTCH2基因3′UTR区序列与GenBank中的已知序列（NG_008163.1）通过BLAST程序对比，检出可能存在的基因突变。采用PicTar和TargetScan在线预测软件预测可能与NOTCH2基因3′UTR区域结合的微小RNA (miRNA)，分析NOTCH2基因3′UTR区域突变对miRNA调控NOTCH2基因表达的影响。结果 检测出NOTCH2基因3′UTR区域1个新发突变和5个已报道的单核苷酸多态性（SNP），分别为2 672位（157020T>G）、rs368873082（156595C>T）、rs835576（156691A>G）、rs3795664（156937C>T）、rs699779（156967T>C）和rs699780（156836T>C）；5个已报道的SNP在TOF组与CON组中的等位基因频率差异均无统计学意义(P>0.05)。预测结果显示34种miRNA能与NOTCH2基因3′UTR区域结合，该突变并不位于34种miRNA与NOTCH2 3′UTR的结合区域。结论 NOTCH2基因3′UTR区域存在新突变157020T>G，该突变有可能通过空间构象影响miRNA与NOTCH2基因3′UTR区的结合效率，进而影响NOTCH2基因的正常表达；而该区域的5个SNP与TOF易感性无明显关联。

关键词: 法洛四联症, NOTCH2基因, 3′UTR, 基因突变, 微小RNA, 单核苷酸多态性

Abstract:

Objective To screen mutations in 3′UTR region of NOTCH2 gene of children with tetralogy of Fallot disease (TOF) and explore possible mechanisms that how mutations in this region affect the expression of NOTCH2 gene. Methods One hundred and fifty-two children confirmed with TOF were selected as subjects (TOF group), and another 107 healthy children were chosen as controls (CON group). PCR was employed to amplify 3′UTR region of NOTCH2 gene. Direct forward and reverse sequencing was performed on the PCR products. The sequence of 3′UTR region of NOTCH2 gene was compared with the known sequence (NG_008163.1) in GenBank by the BLAST program and possible mutations were screened. Online software PicTar and TargetScan were used to predict miRNA which could bind to 3′UTR region of NOTCH2 gene. The effects of mutations in 3′UTR region of NOTCH2 gene on the expression of miRNA-regulated NOTCH2 gene were analyzed. Results A new mutation and 5 reported SNPs in 3′UTR region of NOTCH2 gene, i.e. 2 672(157020T>G), rs368873082(156595C>T), rs835576(156691A>G), rs3795664(156937C>T), rs699779(156967T>C), and rs699780(156836T>C) were detected. The differences of the frequencies of alleles of 5 reported SNPs of the TOF group and CON group were not statistically significant (P>0.05). Results of prediction showed that there were 34 miRNAs that might bind to 3′UTR region of NOTCH2 gene and this mutation did not locate in binding regions of 34 miRNAs and 3′UTR region of NOTCH2 gene. Conclusion A new mutation 157020T>G exists in 3′UTR region of NOTCH2 gene. This mutation may affect the binding efficiency of miRNA and 3′UTR region of NOTCH2 gene by spatial configuration changes, and further affect the normal expression of NOTCH2 gene. Five SNPs in 3′UTR region of NOTCH2 gene are not significantly correlated with the susceptibility of TOF.

Key words: tetralogy of Fallot disease, NOTCH2 gene, 3′UTR, gene mutation, microRNA, single nucleotide polymorphism