Development of a 13-locus PCR multiplex system for paternity testing
Development of a 13-locus PCR multiplex system for paternity testing
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DOI：
10.1007/s00414-003-0420-5
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发表时间：
2004-02-01
影响因子：
2.1
通讯作者：
Lederer, T
中科院分区：
文献类型：
--
作者：
Schlenk, J;Seidl, S;Lederer, T
在这项研究中，描述了适合德国亲子鉴定最新需求的 13 位点多重 PCR 系统的开发。为此，扩展了现有的多重 PCR 系统，该系统允许同时扩增八个不同的 STR 基因座以及性别特异性基因座釉原蛋白（genRESMPX-2，Serac，德国）。虽然一些引物取自底层多重系统，但为 STR 基因座 D19S433、TPOX、TH01、D16S539、D5S818、D2S1338 和 FGA 选择了合适的引物序列。导致片段大小可能重叠的基因座引物用荧光染料 6-FAM、JOE 和 NED 标记。优化反应条件，例如退火温度、引物浓度和聚合酶或缓冲液条件，以获得各种样品来源的稳健扩增和可重复的基因型分析。单一来源样本的完整 DNA 谱可从低至 120 pg 的模板 DNA 中可靠地分型，这表明该系统也可用于法医案例分析。新的多重 PCR 系统的平均排除机会 (MEC) 为 99.9989%，辨别力 (P-D) 约为 1x10(14)（白种人），为法医 DNA 分析提供了重要且灵敏的系统。在这些研究的基础上，Serac（巴特洪堡，德国，genRESMPX-3）现已提供商业套件系统。
In this study the development of a 13-locus multiplex-PCR system fitting the updated demands for paternity testing in Germany is described. For this purpose an existing multiplex PCR system that allows the simultaneous amplification of eight different STR loci together with the sex-specific locus amelogenin (genRESMPX-2, Serac, Germany) was extended. Whereas some of the primers were taken from the underlying multiplex system, suitable primer sequences were chosen for the STR loci D19S433, TPOX, TH01, D16S539, D5S818, D2S1338 and FGA. Primers of loci resulting in potentially overlapping fragment sizes were labelled with the fluorescent dyes 6-FAM, JOE and NED. Reaction conditions, such as annealing temperature, concentrations of primers and polymerase or buffer conditions were optimised to obtain a robust amplification and reproducible genotype analysis for various sample sources. Full DNA profiles from single source samples were reliably typed from template DNA amounts of as low as 120 pg, suggesting a potential use of this system also in forensic casework analysis. With a mean exclusion chance (MEC) of 99.9989% and a power of discrimination (P-D) of about 1x10(14) (Caucasians), the new multiplex PCR system provides a significant and sensitive system for forensic DNA analysis. On the basis of these studies, a commercial kit system is now provided by Serac (Bad Homburg, Germany, genRESMPX-3).