全基因外显子测序技术探寻川崎病易感基因的研究

发布时间：2018-02-10 13:57

本文关键词： 川崎病全基因外显子测序易感基因 SLC15A1基因　出处：《苏州大学》2016年硕士论文　论文类型：学位论文

【摘要】：目的:本课题将对川崎病(Kawasaki disease,KD)病例进行全外显子高通量测序(Whole Exome Sequencing,WES),在全外显子范围内探寻川崎病新的易感基因位点,并进一步探讨易感基因位点与并发冠状动脉损害(Coronary artery lesion,CAL)的关系。方法:我们选取苏州大学附属儿童医院就诊的再发KD患儿6例,作为对照的健康儿童6例,提取外周血DNA,进行全基因组外显子高通量测序,通过数据统计生物信息分析筛选出候选易感位点,对候选易感位点进行聚合酶连反应(Polymerase chain reaction,PCR)扩增和直接测序来检验外显子测序的准确性,并将测序正确的候选易感位点再在50例KD患儿和49例健康儿童的扩大样本中分析,筛选出病例组和健康对照组中基因频率有显著差异的位点,即得到KD的目标易感基因。在扩大样本的50例KD中再分为19例冠脉损害组和31例无冠脉损害组(Non-Coronary artery lesion,NCAL),分析KD的目标易感基因位点在这两亚组中的等位基因频率是否存在显著差异,以探索KD并发CAL的风险等位基因。结果:我们通过全外显子测序和数据统计生物信息分析,筛选出SPEF2、EYS、SLC15A1、PTPN21、HLA-A这5个基因中的12个SNP位点为KD的候选易感位点,检验这些候选易感位点外显子测序准确性,并在扩大的KD病例和对照组中验证分析,进一步证明了SLC15A1 SNPrs2274828为川崎病的目标易感基因位点。在扩大的KD病例组中,SLC15A1 SNPrs2274828位点的基因型CT和等位基因T分布频率显著高于健康对照组(p0.05),在CAL组SLC15A1 SNPrs2274828位点的基因型CT和等位基因T分布频率显著高于NCAL组(p0.05)。结论:1、利用全基因外显子测序的方法,对再发KD和健康对照进行全外显子捕获和测序,通过对比分析发现SLC15A1可能为KD的易感基因。2、SLC15A1 SNPrs2274828位点等位基因T可能为儿童患KD的危险因素。3、SLC15A1 SNPrs2274828位点等位基因T可能增加KD患儿并发CAL的风险性。
[Abstract]:Objective: in this study, whole Exome sequencingus was performed on the whole exon of Kawasaki disease (KDV) in Kawasaki disease, and a new susceptibility gene site of Kawasaki disease was found in the whole exon region of Kawasaki disease (Kawasaki disease), in order to find out the new susceptibility gene of Kawasaki disease (Kawasaki disease). To explore the relationship between susceptibility gene locus and coronary artery lesioning (CAL). Methods: we selected 6 children with recurrent KD who were admitted to the affiliated Children's Hospital of Suzhou University as control group. The peripheral blood DNA was extracted, the high throughput sequencing of the whole genome exon was carried out, and the candidate susceptible sites were screened by statistical bioinformatics analysis. The candidate susceptible sites were amplified by polymerase chain reaction (PCR) and directly sequenced to verify the accuracy of exon sequencing. The correct candidate susceptible sites were analyzed in the expanded samples of 50 KD children and 49 healthy children. The loci with significant difference in gene frequency between the case group and the healthy control group were screened. The target susceptibility gene of KD was obtained. The target susceptibility gene of KD was divided into 19 coronary artery lesion group and 31 non-coronary artery lesion group. The allele frequency of the target susceptibility gene of KD in these two subgroups was analyzed as follows: (1) the allelic frequency of the target susceptibility gene in the two subgroups was as follows: (1) the allele frequency of the target susceptibility gene in the two subgroups was as follows: 1. Whether there is a significant difference, In order to explore the risk alleles of KD complicated with CAL, we screened out 12 SNP loci of the 5 genes of SPEF2EYSU, SLC15A1, PTPN21, HLA-A, which were candidate susceptibility sites for KD by total exon sequencing and statistical bioinformatics analysis. The exon sequencing accuracy of these candidate susceptibility sites was tested, and the analysis was verified in extended KD cases and controls. It is further proved that SLC15A1 SNPrs2274828 is the target susceptibility gene for Kawasaki disease. The frequency of genotype CT and allele T distribution of SLC15A1 SNPrs2274828 locus in extended KD patients is significantly higher than that in healthy control group, and the gene at SLC15A1 SNPrs2274828 locus in CAL group is significantly higher than that in healthy control group. The frequency of T distribution of type type CT and allele was significantly higher than that of NCAL group (P 0.05). Conclusion\% 1, using the method of whole gene exon sequencing, The whole exon was captured and sequenced for recurrent KD and healthy control. It was found that the allele T at SLC15A1 SNPrs2274828 locus of SLC15A1 may be the risk factor of KD in children. The allele T at SNPrs2274828 locus of SLC15A1 may increase the risk of CAL in KD children.
【学位授予单位】：苏州大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R725.4

【相似文献】