Distribution Of 24 SNP Loci In Henan Han Population And Exploring Genes Related To Intracranial Aneurysms

Background and purposeSingle nucleotide polymorphisms(SNPs)is the most common form of human genetic variations that show a sequence polymorphism caused by a single nucleotide change at the genome level.SNP have attracted widespread attention from forensic geneticists due to its advantages of small amplified fragments,low mutation rate,flexible detection methods,and abuntant phenotypic and ancestral information it contain.Currently,several forensic individual genotyping systems based on bi-allelic or tri-allelic SNPs have been established through capillary electrophoresis(CE)and next-generation sequencing(NGS)technologies.As the polymorphic genetic marker,results obtained from SNP were consistent with STR identification when analyzing pedigree-related and phenotypic-related features,especially in degraded samples or traces researchers can get more information compared to STR.Thus we can apply it in individual identification that STR cannot complete.At present,SNPs related to forensic science are mainly genotyped by the SNaPshot technology,because the genotyping platform ABI 3730-XL sequencer is a very popular device in forensic physical and chemical laboratories.SNPs also can be used to study the genetic factors of complex diseases by comparing the allele frequency difference between a large number of patients and the general population in order to discover the effects of genetic variation in complex diseases.Researches on the genetic factors of complex diseases at domestic and foreign have focused on the screen of single nucleotide polymorphisms of candidate genes,namely by analyzing the candidate genes to study the association between this gene and the disease.Thus we can assure that certain sites are genetic risk factors of disease-associated.Intracranial aneurysm(IAs)is a kind of acquired cerebrovascular disease characterized by limited dilation of intracranial artery and thinning of vascular wall due to the weakness of vascular endothelial layer.The statistical analysis reveals that the general incidence of IAs and annual incidence of subarachnoid hemorrhage(SAH).IAs is 3.2%and 9/10000,respectively,the mortality is as high as 50%,and account for about 5%of the various types of cerebrovascular accidents,but 85%of the causes of SAH are due to the rupture of IAs.Epidemiological and genetic studies shown that complex interactions of genetic and environmental risk factors are invloved in IAs.IAs has the characteristics of strong genetic heterogeneity,low penetrance and multi-gene participation,however,its genetic pattern has not yet been determined.The methods of genetic research on intracranial aneurysms include linkage analysis and candidate gene studies,genome-wide association studies,expression studies and whole-exome sequencing.Although genetic studies have identified some susceptibility genes and loci associated with IAs,there are still more risk factors and pathogenic mechanisms need to be discovered.A number of GWAS studies have been conducted on intracranial aneurysms in Asian and European populations and novel potential genetic risk factors have been identified.However,the conclusions are not consistent between different groups.Therefore,it’s necessary to find genes related to intracranial aneurysms in Chinese Han population through SNPs association analysis.The 24 SNPs selected in this study were located in IAs related genes reported before or suspected related gene regions.The genetic polymorphism distributions and differences of these loci in Henan Han population and IAs patient population were investigated to find the genes which related to IAs.This may be helpful in exploring the pathogenesis of intracranial aneurysms,developing new methods of IAs diagnosis and treatment,as well as predicting the risk.At the same time,the population genetic polymorphism information obtained in this study lays a foundation for the application of these loci in forensic medicine.Methods1.A total of 192 patients with intracranial aneurysms admitted from March2015 to March 2017 were collected in the Interventional Department of the first affiliated Hospital of Zhengzhou University.All patients were diagnosed by clinical examination(history,signs,biochemical tests,MRA/CTA/DSA).192 patients had fasting peripheral blood 2 ml in the morning and placed in EDTA anticoagulant tube.The control group was the 188 blood donors collected at the Red Cross Blood Center of Henan Province.2.A total of 380 peripheral blood genome DNA were extracted using 2ml blood genome DNA extraction kit(Lifefeng biotechnology Company,Shanghai)and the concentration and purity of the extracted DNA were measured using a UV spectrophotometer.3.The candidate genes associated with intracranial aneurysms were selected from the literature and the higher MAF of SNPs loci were selected from the candidate genes.Then the fluorescent labeling complex typing system was constructed by using SNaPshot method.The 24 of 100-400bp fragments containing 24 sites were amplified by multiplex PCR reaction.The purified PCR products were amplified by SNaPshot multiplex single base extension reaction.Finally,the ABI3730XL sequencer was used to type the product.4.Calculate the forensic-related parameters and through statistical analysis to analyze the difference in allele frequency between the case group and the control group to determine the effect of genetic variation.Results1.The distribution of the genotype frequencies of 24 SNP loci were consistent with Hardy Weinberg’s equilibrium in Henan Han population.2.The cumulative personal identification ratewas 0.999998,the cumulative probability of exclusion was 0.968814 and the cumulative matching probability was7.875×10~-1010 of the 24 SNP loci in Henan Han population.3.The distribution of the genotype frequencies of the three loci that is rs1072737(χ~2=6.325,Р=0.042),rs17576(χ~2=6.778,Р=0.034)and rs1800255(χ~2=14.387,Р=0.001)between the IA’s group and the normal group showed a sigificant difference(p<0.05)in Henan Han population.4.The distribution of the allele frequenciesrs of the two loci thar is rs2275271(OR=1.343,95%CI=1.007~1.791,Р=0.044),rs180025(OR=1.756, 95%CI=1.225~2.516,Р=0.002)between the IA’s group and the normal group showed a sigificant difference(p<0.05,OR>1,95%CI>1)in Henan Han population.Conclusion1.The selected 24 SNP have application value in individual identification and paternity testing in forensic medicine.2.The polymorphisms of rs17576,rs1800255 and rs1072737 in Henan Han population are associated with intracranial aneurysms.3.The T allele of rs2275271 and the G allele of rs1800255 in Henan Han population may be the genetic risk factors for intracranial aneurysms,which may increase the risk of the disease.