Ricci U, Sani I, Guarducci S, Biondi C, Pelagatti S, Lazzerini V, Brusaferri A, Lapini M, Andreucci E, Giunti L, Giovannucci Uzielli M L
Department of Pediatrics, University of Florence, Italy.
Electrophoresis. 2000 Nov;21(17):3564-70. doi: 10.1002/1522-2683(200011)21:17<3564::AID-ELPS3564>3.0.CO;2-O.
We used an infrared (IR) automated fluorescence monolaser sequencer for the analysis of 13 autosomal short tandem repeat (STR) systems (TPOX, D3S1358, FGA, CSF1PO, D5S818, D7S820, D8S1179, TH01, vWA, D13S317, D16S359, D18S51, D21S11) and the X-Y homologous gene amelogenin system. These two systems represent the core of the combined DNA index systems (CODIS). Four independent multiplex reactions, based on the polymerase chain reaction (PCR) technique and on the direct labeling of the forward primer of every primer pair, with a new molecule (IRDye800), were set up, permitting the exact characterization of the alleles by comparison with ladders of specific sequenced alleles. This is the first report of the whole analysis of the STRs of the CODIS core using an IR automated DNA sequencer. The protocol was used to solve paternity/maternity tests and for population studies. The electrophoretic system also proved useful for the correct typing of those loci differing in size by only 2 bp. A sensibility study demonstrated that the test can detect an average of 10 pg of undegraded human DNA. We also performed a preliminary study analyzing some forensic samples and mixed stains, which suggested the usefulness of using this analytical system for human identification as well as for forensic purposes.
我们使用红外（IR）自动荧光单激光测序仪分析13个常染色体短串联重复序列（STR）系统（TPOX、D3S1358、FGA、CSF1PO、D5S818、D7S820、D8S1179、TH01、vWA、D13S317、D16S359、D18S51、D21S11）以及X-Y同源基因牙釉蛋白系统。这两个系统代表了联合DNA索引系统（CODIS）的核心。基于聚合酶链反应（PCR）技术以及对每个引物对的正向引物进行直接标记，使用一种新分子（IRDye800），建立了四个独立的多重反应，通过与特定测序等位基因的阶梯进行比较，能够准确鉴定等位基因。这是首次使用IR自动DNA测序仪对CODIS核心的STR进行全面分析的报告。该方案用于解决亲子鉴定/亲权鉴定测试以及群体研究。电泳系统对于正确分型那些大小仅相差2 bp的位点也证明是有用的。敏感性研究表明，该测试平均能够检测到10 pg未降解的人类DNA。我们还进行了一项初步研究，分析了一些法医样本和混合污渍，结果表明使用该分析系统进行人类身份鉴定以及法医目的是有用的。
