两个锁骨颅骨发育不全综合征家系的遗传学分析

发布时间：2018-02-14 01:02

本文关键词： 锁骨颅骨发育不全综合征 RUNX2基因基因突变拷贝数改变　出处：《中南大学》2010年硕士论文　论文类型：学位论文

【摘要】： 背景：锁骨颅骨发育不全(Cleidocranial Dysplasia,CCD)是一种罕见的遗传性骨骼系统疾病(MIM 119600),出生发病率为1：100,000。该综合征多为常染色体显性遗传方式,其致病基因定位于染色体6p21。以往研究表明成骨细胞特异性转录因子(RUNX2/CBFA1)的杂合突变、基因插入、缺失等是造成CCD的重要原因。此外,该病具有极强的外显率和明显的家族聚集性,且性别间无明显差异。典型的CCD临床表现包括：锁骨发育不良或无锁骨,囟门和颅缝增宽、延迟闭合或不闭合,身材矮小,出牙晚和恒牙数目增多等骨骼异常。这些骨骼异常所造成的身体畸形给患者带来了沉重的生活压力和心理负担。目的：通过对收集到的两个锁骨颅骨发育不全家系的RUNX2基因突变研究,进一步探索该病的分子发病机制。方法：对临床收集到的两个锁骨颅骨发育不全综合征家系行外周血基因组DNA的提取,利用聚合酶链式反应,DNA直接测序检测突变。对检测到的突变用DHPLC方法在健康人群中进行筛查。对未发现RUNX2基因位点突变的患者行荧光原位杂交检测及全基因组拷贝数检测。结果：家系一中每位CCD患者均存在RUNX2基因c.507delC的杂合突变,从而使171位氨基酸的密码子CTG移码变成TGA而提前终止,改变了RUNX2基因中runt功能域的氨基酸序列。家系二中患者DNA测序未发现RUNX2基因突变,后经包含RUNX2基因的BAC探针荧光原位杂交以及全基因组拷贝数分析显示该患者存在包含RUNX2基因在内的3.5Mb大片段杂合缺失。结论：本研究在家系一患者中检测到的杂合点突变c.507delC造成RUNX2基因编码氨基酸发生移码改变而提前终止。在家系二患者中检测到的3.5Mb大片段杂合缺失包含了整个RUNX2基因,使得RUNX2编码蛋白质的功能完全缺乏。这两种基因组改变分别是引起两家系中CCD患者患病的原因。这两种突变类型是在锁骨颅骨发育不全患者中首次被检测到的RUNX2基因新突变。两种新突变类型的确立进一步扩展了CCD的突变谱,将会对CCD产前诊断和植入前诊断产生积极的影响。
[Abstract]:Background: Cleidocranial Dysplasia (CCDD) is a rare hereditary skeletal system disease with a birth incidence of 1: 100000.This syndrome is mostly autosomal dominant. Previous studies have shown that heterozygosity, gene insertion and deletion of osteoblast specific transcription factor RUNX2 / CBFA1 are important causes of CCD. The typical clinical manifestations of CCD include hypoplasia of clavicle or no clavicle, widening of fontanelle and cranial suture, delayed closure or unclosure, short stature. The abnormal bones, such as late tooth emergence and increasing number of permanent teeth, bring heavy life pressure and psychological burden to the patients due to the body deformities caused by these abnormal bones. Objective: to investigate the molecular pathogenesis of RUNX2 gene mutation in two clavicular cranial dysplasia families. Methods: genomic DNA was extracted from peripheral blood of two families with clavicular cranial dysplasia syndrome. Polymerase chain reaction (PCR) was used to detect mutations by direct sequencing. The detected mutations were screened by DHPLC method in healthy population. Fluorescence in situ hybridization (Fish) and whole genome copy number (copy number) were performed in patients with no mutation of RUNX2 gene. Results: the heterozygosity mutation of RUNX2 gene c.507delC was found in every CCD patient in the first family, thus the 171-amino acid codon CTG frame shifter was transformed into TGA and terminated earlier. The amino acid sequence of runt functional domain in RUNX2 gene was changed. No mutation of RUNX2 gene was found in the DNA sequence of family 2 patients. Fluorescence in situ hybridization with BAC probe containing RUNX2 gene and whole genome copy number analysis showed that the patient had 3.5Mb heterozygosity deletion including RUNX2 gene. Conclusion: the heterozygous point mutation c.507delC detected in this study caused the change of coding amino acids of the RUNX2 gene and terminated early. The 3.5Mb heterozygosity detected in the patient of the second home line contained the entire RUNX2 gene. These two genomes changes are the cause of the disease of CCD patients in two families. These two mutation types are the first detected RUNX2 in patients with clavicular cranial dysplasia. The establishment of two new mutation types further expanded the mutation spectrum of CCD. It will have a positive impact on CCD prenatal diagnosis and pre-implantation diagnosis.
【学位授予单位】：中南大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R681.1;R394

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