云南地区不明原因猝死相关基因突变检测

发布时间：2018-02-02 19:52

本文关键词： 云南猝死基因法医学　出处：《昆明医科大学》2017年硕士论文　论文类型：学位论文

【摘要】：[目的]不明原因粹死(sudden unexpected death,SUD)是当前法医学领域研究的热点问题。云南不明原因粹死(yunnan sudden unexpected death,YNSUD)是特指发生在云南一些偏远山区、半山区,具有明显的家族或家庭聚集性、空间聚集性和时间聚集性等特点的猝死。此外,在云南省各地尚存在无上述聚集性特点的散发性SUD。上述云南地区SUD因病因不明、无有效防治措施,已成为医学、法医学的难点问题。本研究拟检测SUD、YNSUD的相关基因及基因突变,以探索其死亡原因及死亡机制。[方法]1.样本的选取:选取1984年～2015年间昆明医科大学法医学院所做的发生于云南地区的经法医病理解剖、组织学检验及常规毒药物检验等全面检验后仍不能明确死亡原因的猝死案例,共95例。2.对90例散发性SUD案例、3例YNSUD案例的KCNQ1-Exon3第572-592碱基、KCNQ1-Exon6 第 805-905 碱基、KCNH2-Exon7 第 1681-1902 碱基、KCNH2-Exon9第 2398+5 碱基、SCN5A-Exon3 第 283-362 碱基、SCN5A-Exon28 第 5291-6016 碱基利用软件PrimerPremier 5.0设计引物、PCR扩增、DNA直接测序。测序结果通过NCBI数据库比对筛查出突变位点并对错义突变位点进行蛋白质功能预测,其蛋白质预测结果用pspe表示。3.对2例YNSUD案例(互为母女)及其家系样本1例(祖父),进行全基因组测序。对结果进行家系分析。[结果]1.猝死相关基因突变位点筛查结果:(1)90例散发性SUD案例中有28例样本检测出猝死相关的30个突变位点。KCNQ1基因3号外显子c.576GT(p.R192L)筛查出1例错义突变样本,其pspe=0.982。KCNH2基因7号外显子筛查出4个位点的杂合突变,1例样本筛查出c.1789TA(p.Y597N)杂合突变,其psp e=0.994;1例样本筛查出c.1769GA(p.G590D)杂合突变,其pspe=0.559;19例样本筛查出c.1801GA(p.G601S)杂合突变,其pspe=0.125;10例样本筛查出 c.1800CA(p.S600R)杂合突变,其 pspe=0.03。KCNH2 基因 7 号外显子 c.1765CT、SCN5A 基因 28 号外显子 c.5295TC、SCN5A 基因 28 号外显子 c.5447CT各筛查出1例同义突变。(2)3例YNSUD案例中,KCNH2基因7号外显子筛查出 c.1801GA(p.G601S)1 例杂合突变样本,其 pspe=0.125。2.全基因组测序筛查出与SUD相关的OBSCN、TTN基因的共5个突变位点,其突变位点分别为1号染色体OBSCN基因c.228548197GA(密码子变化:cGt/c At),c.228550426CT(密码子变化:Ccc/Tcc),c.228564884GA(密码子变化:cGc/cAc);2 号染色体 TTN 基因 c.179558366AG(密码子变化:Att/Gtt),c.179582537CG(密码子变化:gCg/gAg)。[结论]1.SUD、YNSUD案例中,筛查出五个突变位点,分别是KCNQ1基因576位置,KCNH2 基因 1789、1769、1800、1801 位置。2.通过对筛查出的错义突变位点进行蛋白功能预测发现,KCNQ1基因3号外显子 c.576GT(p.R192L)和 KCNH2 基因 7 号外显子 c.1789TA(p.Y597N)的突变较可能导致对应离子通道蛋白功能改变,可能与SUD相关。3.通过全基因组测序发现与猝死相关的OBSCN基因c.228548197GA、c.228550426CT、c.228564884GA 和 TTN 基因 c.179558366AG、c.179582537CG 的错义突变。4.SUD、YNSUD可能与相关基因突变有关。
[Abstract]:[Objective] to die of unexpected death of unknown cause. Sud) is a hot issue in the field of forensic medicine. Yunnan unknown cause death sudden unexpected death. YNSUD) is a sudden death that occurs in some remote mountainous and semi-mountainous areas of Yunnan Province with obvious family or family agglomeration spatial aggregation and temporal agglomeration. There are sporadic SUDs in all parts of Yunnan Province without the above characteristics of aggregation. The SUD in Yunnan area has become a medicine because of its unknown etiology and no effective prevention and treatment measures. In order to explore the cause of death and the mechanism of death, this study intends to detect the related genes and gene mutations of SUDYNSUD. [Methods: 1. Sample selection: selected from 1984 to 2015 by the Institute of Forensic Medicine of Kunming Medical University, which occurred in Yunnan through forensic pathology anatomy. 2. There were 95 cases of sudden death after histological examination and routine drug test. 90 cases were sporadic SUD. KCNQ1-Exon6 nucleotide 805-905 of KCNQ1-Exon3 572-592 in 3 cases of YNSUD. KCNH2-Exon7 1681-1902 base KCNH2-Exon9 23985. SCN5A-Exon3 283-362 bases. The 5291-6016 base of SCN5A-Exon28 was amplified by PrimerPremier 5.0 primer. DNA was sequenced directly. The results of sequencing were compared with NCBI database to screen mutation sites and predict the protein function of missense mutation sites. The results of protein prediction were expressed as pspe. 3. Two cases of YNSUD (mother and daughter) and one case of family (grandfather) were sequenced. The results were analyzed by family analysis. [Results: 1. Screening for mutation sites of sudden death related genes: 1. Thirty mutation sites associated with sudden death were detected in 28 of 90 sporadic SUD cases. Exon 3 of the KCNQ1 gene, c. 576GTp.R1921, was detected. A sample of missense mutation was screened. The pspe=0.982.KCNH2 gene exon 7 was screened for 4 loci heterozygous mutations in a sample of c. 1789 TAP. Y597N). Psp eosin 0.994; The heterozygous mutation of C. 1769 GAP. G590D was screened out in one case, and its psper was 0.559; The heterozygous mutation of c. 1801GAP. G601S was screened out in 19 samples, and its pspeChina was 0.125; The heterozygosity mutation of c. 1800 CAP. S600R was screened in 10 samples, and its pspe=0.03.KCNH2 gene exon 7 c. 1765CT. SCN5A gene exon 28 c. 5295TC. Exon 28 of SCN5A gene, c. 5447CT, was used to screen 1 case of synonymous mutation and 3 cases of YNSUD. Exon 7 of KCNH2 gene was used to screen a heterozygous mutation sample of c. 1801GAP. G601SU. The whole genome was sequenced to screen five mutation sites of OBSCN TTN gene associated with SUD. The mutation sites were the OBSCN gene c.228548197GAon of chromosome 1 (codon variation: C GtP / c At). C. 228550426CT. Chromosome 2: TTN gene c.179558366AG. C. 179582537 CG' [Conclusions: 1. In the case of sud YNSUD, five mutation loci were screened, which were KCNQ1 gene 576, KCNH2 1789 and 1769 / 1800. 1801 position. 2. Protein function prediction was carried out by detecting the missense mutation sites. Exon 3 of KCNQ1 gene c. 576GTp.R19L) and exon 7 of KCNH2 gene c. 1789 TAA p.Y597N). The mutation may result in the functional change of the corresponding ion channel protein. OBSCN gene c. 228548197 GAN c. 228550426CT, which may be associated with SUD. C. 228564884GA and TTN gene c.179558366AGN c.179582537CG missense mutation .4.SUD. YNSUD may be associated with related gene mutations.
【学位授予单位】：昆明医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：D919.4

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