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浙江大学学报（医学版）
浙江大学学报（医学版）  2019, Vol. 48 Issue (4): 373-377    DOI: 10.3785/j.issn.1008-9292.2019.08.04
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CLN6基因复合杂合突变导致神经元蜡样脂褐质沉积症一家系遗传学研究
楼铁1,2(),黄颖之1,董旻岳1,*()
1. 浙江大学医学院附属妇产科医院生殖遗传科 生殖遗传教育部重点实验室, 浙江 杭州 310006
2. 杭州市江干区人民医院妇产科, 浙江 杭州 310021
Genetic study of a family of neuronal ceroid lipofuscinosis caused by a heterozygous mutation of CLN6 gene
LOU Tie1,2(),HUANG Yingzhi1,DONG Minyue1,*()
1. Key Laboratory of Reproductive Genetics, Ministry of Education, Department of Reproductive Genetics, Women's Hospital, Zhejiang University School of Medicine, Hangzhou 310006, China
2. Department of Gynecology and Obstetrics, Jianggan District People's Hospital, Hangzhou 310021, China
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摘要：

目的: 探讨一个常染色体隐性遗传神经元蜡样脂褐质沉积症（NCL）家系的遗传学原因。方法: 应用目标区捕获高通量靶向测序对先证者进行候选基因突变筛查，并通过PCR测序在先证者及其父母中对突变位点进行验证；RT-PCR及TA克隆测序考察上述两种突变是否影响剪接。结果: 测序结果显示先证者存在CLN6：c.486+2T>C和c.486+4A>T复合杂合突变，分别来自父母双方。RT-PCR及TA克隆测序提示两种突变均可导致两种异常剪接。结论: CLN6：c.486+2T>C和c.486+4A>T复合杂合突变很可能为该NCL家系患者的遗传学病因，新突变基因丰富了CLN6基因突变谱。
关键词： 神经元蜡样质脂褐质沉积病/遗传学;  病因学;  基因;  突变;  剪接体;  聚合酶链反应    
Abstract:

Objective: To analyze the genetic cause of a family with autosomal recessive neuronal ceroid lipofuscinoses (NCL). Methods: The proband was screened for mutations within the coding region of the candidate genes through high-throughput targeted sequencing. Potential causative mutations were verified by PCR and Sanger sequencing in the proband and his parents. RT-PCR and TA clone sequencing were performed to investigate whether the mRNAs were abnormally spliced. Results: The sequencing results revealed compound heterozygous mutations of CLN6:c.486+2T>C and c.486+4A>T, which were respectively inherited from his parents. RT-PCR and TA cloning sequencing suggested that the mRNAs were abnormally spliced in two forms due to both mutations. Conclusions: The compound heterozygous mutations of CLN6:c.486+2T>C and c.486+4A>T are possibly the genetic causes of the NCL family. Detection of the novel mutation has extended mutation spectrum of CLN6.
Key words: Neuronal ceroid-lipofuscinoses/genetics    Etiology    Genes    Mutation    Spliceosomes    Polymerase chain reaction
收稿日期: 2019-03-28 出版日期: 2019-10-30
:  R394.3  
基金资助: 浙江省重点研发计划(2019C03025)
通讯作者: 董旻岳     E-mail: tiel-2004@163.com;dongmy@zju.edu.cn
作者简介: 楼铁(1978—), 女, 硕士, 主治医师, 主要从事妇产科学研究; E-mail: tiel-2004@163.com; https://orcid.org/0000-0002-1069-3803
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引用本文:

楼铁,黄颖之,董旻岳. CLN6基因复合杂合突变导致神经元蜡样脂褐质沉积症一家系遗传学研究[J]. 浙江大学学报（医学版）, 2019, 48(4): 373-377.

LOU Tie,HUANG Yingzhi,DONG Minyue. Genetic study of a family of neuronal ceroid lipofuscinosis caused by a heterozygous mutation of CLN6 gene. J Zhejiang Univ (Med Sci), 2019, 48(4): 373-377.

链接本文:

http://www.zjujournals.com/med/CN/10.3785/j.issn.1008-9292.2019.08.04    或    http://www.zjujournals.com/med/CN/Y2019/V48/I4/373

引物 引物序列(5′-3′) 片段长度
(bp)*
*此为标准参考序列长度，实际cDNA片段大小可随剪接异常而变化.
CLN6-4F GTGACATTCCAGCTGGAAGCGT 467
CLN6-4R GAACACTTGAGCATCCTAGCTTG
CLN6-mrE3-F CTTCCACATGGCCTACAACGTC 300
CLN6-mrE6-R GCAGTAAAGCAGCCGCTGAAGT
CLN6-mrE3_E4-F TTCTCTTGCTCAAGCTCATCGAG 300
CLN6-mrE5_E6-R GGATGTACCACATGCAGTGACC
表 1  引物序列
图 1  先证者及其父母 CLN6 基因的Sanger测序结果
图 2  先证者cDNA TA克隆测序结果
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