疑难亲缘鉴定中非CODIS STR和X-SNP的应用价值
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摘要
目的:通过对26个常染色体非CODIS-STR基因座在中国汉族人群中的多态性调查和建立67个X-SNP位点的MALDI-TOF MS检测技术,为疑难亲权鉴定案件如突变案例、特殊排除案例、祖母与孙女的亲缘关系鉴定、同父异母姐妹关系的鉴定等提供鉴定思路和解决方法。
方法:采用AGCU 21+1试剂盒调查20个常染色体STR基因座在中国汉族人群的多态性,计算其遗传学参数;建立多重复合扩增体系调查6个常染色体STR基因座中国汉族人群的多态性,计算其遗传学参数。并在实际案例中考察这27个STR基因座的应用价值。建立MALDI-TOF MS技术检测67个X-SNP位点的方法,并调查67个X-SNP位点在中国汉族人群的多态性、群体遗传学参数和连锁情况等。
结果:26个常染色体STR基因座在中国汉族人群中具有良好的多态性和理想的遗传学参数,其在研究人群中频率分布均符合Hardy-Weinberg平衡。三联体累积非父排除率(CPE)为0.999999996,二联体累积非父排除率为0.999985,累积个人识别能力(CDP)为0.99999999999999999999999997。67个X-SNP位点中66个位点符合Hardy-Weinberge平衡,有3个位点的最低等位基因频率(MAF)低于0.1,其余64个位点均具有高信息量,并且有8组SNP存在紧密连锁,通过筛选得到56个X-SNP位点。女性群体中的CDP达到0.999999999999999999999985,在男性群体的CDP为0.999999999999977,在三联体的CPE为0.999999999987,在二联体的CPE为0.999999765。
结论:26个常染色体非CODIS STR基因座具有良好的遗传多态性,能有效运用于法医学个人识别和亲子鉴定,并且能够在疑难亲权案例鉴定中发挥重要的补充作用;MALDI-TOF MS技术对X-SNP位点的分型准确可靠,通过筛选得到的56个X-SNP位点具有良好的遗传多态性,累积个体识别能力和非父排除率已能满足法医学应用的要求,可为特殊亲缘鉴定案件提供解决手段。
Objective: Through investigating the genetic polymorphisms 26 autosomal non-CODIS STR loci in Chinese Han Population and establishing the MALDI-TOF MS platform for 67 SNPs on X-Chromosome to provide methods for special cases such as mutational cases、special excluded cases、the judgement of relationships of grandmother to granddaughter and half-sib sisters with the same father.
Methods: The polymorphisms of 21 autosomal non-CODIS STR loci in Chinese Han population were detected by the AGCU 21+1 STR kit .The genetic parameters of them were calculated; The multiplex-PCR system of the six autosomal non-CODIS STR loci was established to detect their polymorphisms in Chinese Han population, and genetic parameters of them were calculated. Application Value of 26 STR loci was investigated through applying in practical cases . Using The MALDI-TOF technique to detect the 67 SNPs on X-Chromosome, then to investigate their polymorphisms in Chinese Han population、genetic parameters and linkage information.
Results :The 26 non-CODIS STR loci were polymorphic in Han population with ideal genetic parameters. Allele frequency distributions of the 26 non-CODIS STR loci did not statistically deviated from the laws of Hardy-Weinberg equilibration. The cumulative exclusionary power(CPE) of trios-testing was 0.999999996, the CPE of duo-testing was 0.99998467, and the cumulated discrimination power(CDP)was 0.99999999999999999999999997. There were 66 X-SNPs following Hardy-Weinberg equilibration in all of 67 sites, and 3 SNPs’low allele frequency were below 0.1, and 8 pairs SNPs existed close linkage. Finally we got 56 X-SNP with high information. The CDP in females reach 0.999999999999999999999985,The CDP in males reach 0.999999999999977,the CPE of trios-testing was 0.999999999987, the CPE of duo-testing was 0.999999765。
Conclusion : The 26 non CODIS STR loci are highly polymorphic. They can be effectively used in personal identification and paternity testing. They can also be used as important supplementary loci in difficult paternity testing cases. The genotyping of X-SNPs by MALDI-TOF MS technology was accurate and reliable. The selected 56 X-SNPs are highly polymorphic, their CDP and CPE can been able to satisfy the requirement of forensic science, and provide new methods for special paternity testing.
方法:采用AGCU 21+1试剂盒调查20个常染色体STR基因座在中国汉族人群的多态性,计算其遗传学参数;建立多重复合扩增体系调查6个常染色体STR基因座中国汉族人群的多态性,计算其遗传学参数。并在实际案例中考察这27个STR基因座的应用价值。建立MALDI-TOF MS技术检测67个X-SNP位点的方法,并调查67个X-SNP位点在中国汉族人群的多态性、群体遗传学参数和连锁情况等。
结果:26个常染色体STR基因座在中国汉族人群中具有良好的多态性和理想的遗传学参数,其在研究人群中频率分布均符合Hardy-Weinberg平衡。三联体累积非父排除率(CPE)为0.999999996,二联体累积非父排除率为0.999985,累积个人识别能力(CDP)为0.99999999999999999999999997。67个X-SNP位点中66个位点符合Hardy-Weinberge平衡,有3个位点的最低等位基因频率(MAF)低于0.1,其余64个位点均具有高信息量,并且有8组SNP存在紧密连锁,通过筛选得到56个X-SNP位点。女性群体中的CDP达到0.999999999999999999999985,在男性群体的CDP为0.999999999999977,在三联体的CPE为0.999999999987,在二联体的CPE为0.999999765。
结论:26个常染色体非CODIS STR基因座具有良好的遗传多态性,能有效运用于法医学个人识别和亲子鉴定,并且能够在疑难亲权案例鉴定中发挥重要的补充作用;MALDI-TOF MS技术对X-SNP位点的分型准确可靠,通过筛选得到的56个X-SNP位点具有良好的遗传多态性,累积个体识别能力和非父排除率已能满足法医学应用的要求,可为特殊亲缘鉴定案件提供解决手段。
Objective: Through investigating the genetic polymorphisms 26 autosomal non-CODIS STR loci in Chinese Han Population and establishing the MALDI-TOF MS platform for 67 SNPs on X-Chromosome to provide methods for special cases such as mutational cases、special excluded cases、the judgement of relationships of grandmother to granddaughter and half-sib sisters with the same father.
Methods: The polymorphisms of 21 autosomal non-CODIS STR loci in Chinese Han population were detected by the AGCU 21+1 STR kit .The genetic parameters of them were calculated; The multiplex-PCR system of the six autosomal non-CODIS STR loci was established to detect their polymorphisms in Chinese Han population, and genetic parameters of them were calculated. Application Value of 26 STR loci was investigated through applying in practical cases . Using The MALDI-TOF technique to detect the 67 SNPs on X-Chromosome, then to investigate their polymorphisms in Chinese Han population、genetic parameters and linkage information.
Results :The 26 non-CODIS STR loci were polymorphic in Han population with ideal genetic parameters. Allele frequency distributions of the 26 non-CODIS STR loci did not statistically deviated from the laws of Hardy-Weinberg equilibration. The cumulative exclusionary power(CPE) of trios-testing was 0.999999996, the CPE of duo-testing was 0.99998467, and the cumulated discrimination power(CDP)was 0.99999999999999999999999997. There were 66 X-SNPs following Hardy-Weinberg equilibration in all of 67 sites, and 3 SNPs’low allele frequency were below 0.1, and 8 pairs SNPs existed close linkage. Finally we got 56 X-SNP with high information. The CDP in females reach 0.999999999999999999999985,The CDP in males reach 0.999999999999977,the CPE of trios-testing was 0.999999999987, the CPE of duo-testing was 0.999999765。
Conclusion : The 26 non CODIS STR loci are highly polymorphic. They can be effectively used in personal identification and paternity testing. They can also be used as important supplementary loci in difficult paternity testing cases. The genotyping of X-SNPs by MALDI-TOF MS technology was accurate and reliable. The selected 56 X-SNPs are highly polymorphic, their CDP and CPE can been able to satisfy the requirement of forensic science, and provide new methods for special paternity testing.
引文
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[2] Vallone PM, Decker AE, Butler JM. Allele frequencies for 70 autosomal SNP loci with U.S. Caucasian, African American, and Hispanic samples[J]. Forensic Sci Int, 2005, 149:279- 286.
[3]台运春,陆惠玲,刘超,等.15个常染色体STR基因座的突变情况[J].中国法医学最新科研与实践[M],北京:中国人民公安大学出版社,2004:209-211.
[4]赵珍敏,柳燕,林源.IdentifilerTM系统在亲子鉴定中的突变观察和分析[J].法医学杂志.2007.23(4):290-294.
[5] Coble MD, Just RS, O’Callaghan IH, et al. Single nucleotide polymorphisms over the entire mtDNA genome that increase the power of forensic testing in Caucasians[J]. Int J Legal Med,2004,118:137- 146.
[6] Zarrabeitia MT, Mijares V, Riancho JA. Forensic efficiency of microsatellites and single nucleotide polymorphisms on the X chromosome[J]. Int J Legal Med,2007, 121(6):433- 437.
[7] Lander ES. The new genomics: global views of biology [J]. Science, 1996,274(5287):536-539.
[8] Brooks A J. The essence of SNPs. Gene,1999,234:177-186.
[9] Li CT, Li RY. SNP genotyping by multiplex amplification and microarrays assay for forensic application[J]. Forensic Sci Int, 2006,162(1- 3): 74- 79.
[10] Petkovski E, Keyser- Tracqui R, Hienne R. SNPs and MULDI-TOF MS: tools for DNA typing in forensic paternity testing and anthropology[J]. J Foren. Sci, 2005, 50:535- 541.
[11] Beatriz S. SNPs in forensic genetics: a review on SNP typing methodologies[J]. Forensic Sci Int,2005, 154:181- 194.
[12] Kidd KK, Pakstis AJ, Speed WC, et al. Developing a SNP panel for forensic identification of individuals[J]. Forensic Sci Int, 2006,164(1):20- 32.
[13] Dixon LA, Dobbins AE, Pulker HK, et al. Analysis of artificially degraded DNA using STRs and SNPs - - results of a collaborative European (EDNAP) exercise [J].Forensic Sci Int, 2006,164(1):33- 44.
[14] Vallone PM, Decker AE, Butler JM. Allele frequencies for 70 autosomal SNP loci with U.S. Caucasian, African American, and Hispanic samples [J]. Forensic Sci Int, 2005, 149:279- 286.
[15] Sanchez JJ, Borsting C, Hallenberg C, et al. Multiplex PCR and minisequencing of SNPs- A model with 35 Y chromosome SNPs [J]. Forensic Sci Int, 2003, 137:74- 84.
[16] Coble MD, Just RS, O’Callaghan IH, et al. Single nucleotide polymorphisms over the entire mtDNA genome that increase the power of forensic testing in Caucasians [J]. Int J Legal Med, 2004, 118: 137- 146.
[17] Zarrabeitia MT, Mijares V, Riancho JA. Forensic efficiency of microsatellites and single nucleotide polymorphisms on the X chromosome[J]. Int J Legal Med,2007, 121(6):433- 437.
[18] Wang X, Ito S, Sawaguchi A, et al. Analysis of single nucleotide polymorphisms and its application in a disputed paternity case [J]. International Congress Series, 2006,1288:76- 78.
[19] Tomas C,Sanchez JJ,Barbaro A,et al.X-chromosome SNP analyses in 11 human Mediterrannean populations show a high overall genetic homogeneity except in North-west African(Moroccans) [J]. BMC Evol Biol. 2008 Feb 29;8:75.
[20] Pereira V, Tomas C, Amorim A, et al. Study of 25 X-chromosome SNPs in the Portuguese. Forensic Sci Int Genet, 2010,
[21] Walsh PS, Metzger DA, Higuchi R, et al. Chelex 100 as a medium for simple extraction of DNA for PCR-based typing form forensic material[J].Biotechniques,1991,10:506-518.
[22] Sinofiler Kit manual,Applied Biosystems,2007.
[23]无锡中德美联生物技术有限公司.AGCU21+1STR荧光检测试剂盒试用装产品说明书[K].
[24]Promega Biotech Co., Ltd. Powerstates V12.xls.[EB/OL] http://www.promega.com/geneticidtools/.
[25]侯一平主编.法医物证学[M].北京:人民卫生出版社(第2版), 2005:223.
[26] Brinkmann B,Klintschar M, Neuhuber F, et al. Mutation rate in human microsatellites: influence of the structure and length of the tandem repeat[J].Am.J.Hum.Genet,1998,62;1408-1415.
[27]李秋阳,丰伟军,杨庆恩,等.常用STR基因座突变的观察与分析[J].中国法医学杂志,2008,23(6):394-396.
[28]邓志辉,吴国光,程良红,等.13个STR基因座在亲子鉴定案例中基因突变观察[J].中国法医学杂志,2003,18(3):150-153.
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