儿童特发性全身性癫痫易感基因CACNA1H及GABRB3的分析研究

发布时间：2018-06-08 15:53

本文选题：特发性全身性癫痫 + CACNA1H基因　；参考：《广西医科大学》2012年博士论文

【摘要】：第一部分特发性全身性癫痫CACNA1H基因的突变分析目的：探讨T型钙通道CACNA1H基因变异与中国广西汉族儿童特发性全身性癫痫(IGE)的关系。方法：对100例中国广西汉族IGE患儿和100例汉族健康对照者应用聚合酶链式反应(Polymerase Chain Reaction,PCR)方法扩增CACNA1H基因外显子9～11编码区及两侧各50个碱基以上的序列,PCR产物经ABI-PRISM3730全自动DNA测序仪自动测序,然后与NCBI数据库中CACNA1H基因的序列进行比对,了解CACNA1H基因突变情况,利用生物信息学方法预测CACNA1H基因突变对蛋白质功能的影响。结果： 1.在IGE患儿中,发现CACNA1H基因两个新的杂合子突变,1254211GA及1254303AT,均位于第10外显子上。分别导致所编码的氨基酸发生改变,Arg735G1n及Arg766Trp。 2. CACNA1H基因编码第735位及第766位Arg在生物进化过程中处于部分保守区域。 3.用SIFT和PolyPhen程序预测CACNA1H基因点突变,显示Arg766Trp有可能影响蛋白质的功能。 4.共发现CACNA1H基因的14个多态性位点,其中有5个变异在NCBI的SNP数据库中未见收录,在IGE组及对照组中比较了7个SNPs位点的基因型、等位基因及单体型的频率分布,结果显示无显著性差异。结论：T型钙通道CACNA1H基因突变可能与中国广西IGE患儿的发生有关,CACNA1H基因很可能是IGE的易感基因。第二部分GABRB3基因多态性与儿童特发性全身性癫痫相关性研究目的：探讨GABRB3基因多态性与中国广西汉族儿童特发性全身性癫痫(IGE)的关系。方法：以GABRB3基因上5个单核苷酸多态性(SNPs)rs4906902,rs981778,rs2059574,rs890317,rs1426217为遗传标记,运用聚合酶链反应-限制性片段长度多态性及DNA测序法对100例广西汉族IGE患儿及200例汉族健康对照者进行5个SNPs位点的测定,应用SPSS17.0软件比较两组儿童中各SNPs位点基因型频率及等位基因频率,用SHESIS软件进行单体型的构建及分析。结果：GABRB3基因中单个SNPs的基因型频率及等位基因频率在IGE组及对照组之间无显著性差异。GABRB3基因单体型A-A-T-A-G(SNPs顺序：rs4906902,rs981778,rs2059574,rs890317,rs1426217)在IGE组出现的频率明显高于正常对照组,差异有统计学意义。结论：γ一氨基丁酸A型受体亚单位GABRB3基因可能与中国广西IGE患儿相关,GABRB3基因可能是广西IGE患儿的易感基因。
[Abstract]:Part I: mutation analysis of CACNA1H gene in idiopathic generalized epilepsy objective: to investigate the relationship between the variation of CACNA1H gene in T-type calcium channel and IGE in Han Chinese children of Guangxi Han nationality. Methods: 100 cases of Han nationality in Guangxi were studied. PCR products of CACNA1H gene exon 9M11 and 50 bases on both sides were amplified by polymerase chain reaction Polymerase chain reaction (Polymerase chain reaction) PCRmethod. The PCR products were sequenced by ABI-PRISM3730 automatic DNA sequencer. Then the sequence of CACNA1H gene was compared with the sequence of CACNA1H gene in NCBI database to understand the mutation of CACNA1H gene, and the effect of CACNA1H gene mutation on protein function was predicted by bioinformatics. Results: 1. Two new heterozygous mutations of CACNA1H gene, 1254211GA and 1254303AT, were found in children with IGE, both at exon 10. The results showed that the encoded amino acids changed Arg735G1n and Arg766TRP. 2, respectively. CACNA1H gene encodes 735th and 766-bit Arg in a partially conserved region during biological evolution. The point mutation of CACNA1H gene was predicted by sift and PolyPhen program, which indicated that Arg766Trp might affect the function of protein. 4. A total of 14 polymorphic loci of CACNA1H gene were found, 5 of which were not included in the SNP database of NCBI. Genotypes, alleles and haplotypes of 7 SNPs loci were compared in the IGE group and the control group. Conclusion the mutation of CACNA1H gene may be related to the occurrence of IGE in Guangxi, China. The second part of GABRB3 gene polymorphism is associated with idiopathic systemic IGE in children. Objective: to investigate the relationship between GABRB3 gene polymorphism and IGE in Chinese Han nationality children with idiopathic generalized epilepsy. Methods: five single nucleotide polymorphisms (SNPs4906902rs981778 rs2059574rs890317rs1426217) of GABRB3 gene were used as genetic markers. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and DNA sequencing were used to detect 5 SNPs loci in 100 children with IGE in Guangxi Han nationality and 200 healthy controls. SPSS 17.0 software was used to compare the genotype frequency and allele frequency of each SNPs locus between the two groups. Results there was no significant difference in genotype frequency and allele frequency of single SNPs between IGE group and control group. The order of haplotype A-T-A-SNPs of GABRB3 gene was: rs4906902rs981778rs2059574rs890317rs1426217). The rate was significantly higher than that in the normal control group. Conclusion: the GABRB3 gene of 纬 -aminobutyric acid type A receptor subunit may be the susceptible gene of IGE children in Guangxi, China.
【学位授予单位】：广西医科大学
【学位级别】：博士
【学位授予年份】：2012
【分类号】：R742.1

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