title: Optimizing direct amplification of forensic commercial kits for STR determination.
abstract: Direct DNA amplification in forensic genotyping reduces analytical time when large sample sets are being analyzed. The amplification success depends mainly upon two factors: on one hand, the PCR chemistry and, on the other, the type of solid substrate where the samples are deposited. We developed a workflow strategy aiming to optimize times and cost when starting from blood samples spotted onto diverse absorbent substrates. A set of 770 blood samples spotted onto Blood cards, Whatman®3 MM paper, FTA™ Classic cards, and Whatman®Grade 1 was analyzed by a unified working strategy including a low-cost pre-treatment, a PCR amplification volume scale-down, and the use of the 3500 Genetic Analyzer as the analytical platform. Samples were analyzed using three different commercial multiplex STR direct amplification kits. The efficiency of the strategy was evidenced by a higher percentage of high-quality profiles obtained (over 94%), a reduced number of re-injections (average 3.2%), and a reduced amplification failure rate (lower than 5%). Average peak height ratio among different commercial kits was 0.91, and the intra-locus balance showed values ranging from 0.92 to 0.94. A comparison with previously reported results was performed demonstrating the efficiency of the proposed modifications. The protocol described herein showed high performance, producing optimal quality profiles, and being both time and cost effective.
authors.affiliation: Universidad de Buenos Aires, Facultad de Farmacia y Bioquímica, Departamento de Microbiología, Inmunología y Biotecnología, Cátedra de Genética Forense y Servicio de Huellas Digitales Genéticas, Junín 956 C1113AAD, Buenos Aires, Argentina; CONICET - Consejo Nacional de Investigaciones Científicas y Tecnológicas, C1033AAJ, Buenos Aires, Argentina. Electronic address: mcaputo@ffyb.uba.ar.
authors.forename: M
authors.initials: M
authors.lastname: Caputo
authors.affiliation: CONICET - Consejo Nacional de Investigaciones Científicas y Tecnológicas, C1033AAJ, Buenos Aires, Argentina. Electronic address: cebobillo@yahoo.com.
authors.forename: M C
authors.initials: MC
authors.lastname: Bobillo
authors.affiliation: Universidad de Buenos Aires, Facultad de Farmacia y Bioquímica, Departamento de Microbiología, Inmunología y Biotecnología, Cátedra de Genética Forense y Servicio de Huellas Digitales Genéticas, Junín 956 C1113AAD, Buenos Aires, Argentina; CONICET - Consejo Nacional de Investigaciones Científicas y Tecnológicas, C1033AAJ, Buenos Aires, Argentina. Electronic address: asala@ffyb.uba.ar.
authors.forename: A
authors.initials: A
authors.lastname: Sala
authors.affiliation: Universidad de Buenos Aires, Facultad de Farmacia y Bioquímica, Departamento de Microbiología, Inmunología y Biotecnología, Cátedra de Genética Forense y Servicio de Huellas Digitales Genéticas, Junín 956 C1113AAD, Buenos Aires, Argentina; CONICET - Consejo Nacional de Investigaciones Científicas y Tecnológicas, C1033AAJ, Buenos Aires, Argentina. Electronic address: dcorach@yahoo.com.
authors.forename: D
authors.initials: D
authors.lastname: Corach
mesh_terms: D004247:DNA / Q000097:blood; D016172:DNA Fingerprinting / Q000379:methods*; D006801:Humans; D018895:Microsatellite Repeats; D060885:Multiplex Polymerase Chain Reaction; D013048:Specimen Handling / Q000295:instrumentation*; D057188:Workflow
chemical_list: D004247:DNA
keywords: Blood stains; Commercial direct DNA amplification kits; High-throughput data banking