常染色体21个SNPs多态性分型方法研究

Study on genotyping method of a 21-locus autosomal SNP multiplex

目的建立常染色体21个SNPs的多态性分型方法。方法采用荧光标记公用引物和等位基因特异性引物原理设计SNP复合扩增引物体系,对45个备选SNP位点筛选,选出21个及性别Amelogenin构成复合扩增体系。PCR产物经3130XL型电泳仪电泳分离,GeneMaperTM3.0数据分析软件分析结果。同时随机选取6份样品,使用测序方法对SNP分型并进行测序验证。结果应用本研究建立的复合扩增体系扩增样品,产物经毛细管电泳后,每个SNPs均可正确判定基因型。随机选取6份样品SNPs位点测序结果显示,荧光标记SNPs复合扩增分型与直接测序结果完全一致。结论本研究建立的荧光标记公有引物特异性片段常染色体21个SNPs复合扩增方法是SNP多态性分析的一种有效方法,并有助于解决SNP分型识别能力、效率、通量和高成本的问题。

Objective To establish the genotyping method of a 21-locus autosomal SNP multiplex.Methods The strategies of universal reporter primer（URP） and allele-specific amplification（ASA） were adopted to build the SNP multiplex system.After screened from 45 autosomal SNPs,Twenty-one SNPs were used to construct the multiplex analysis system with the amelogenin together.The PCR products were separated by 3130XL sequencer and the results were analyzed by GeneMaperTM 3.0.In addition,we have validated the multiplex assay by comparing with the sequencing analysis of 6 samples choosed randomly.Results Using the multiplex system developed by us,all samples were determined the genotype correctly for 21 autosomal SNPs after capillary electrophoresis.Compared with the sequencing analysis of 6 samples choosed randomly for 21 autosomal SNPs,it was consistent with the corresponding results from the multiplex system.Conclusion The multiplex system of 21 autosomal SNPs is an effective method for SNP analysis and is helpful to solve the problems of DP,efficiency,throughput and cost.