02172nam a2200241 4500 393968 393968 20160531d2014 y0pory50 ba eng Microchip analyzer for forensic short tandem repeat typing of single cells Recurso eletrónico final technical report Tao Geng, Richard A. Mathies [Rockville, MD] National Criminal Justice Reference Service (NCJRS) 2014 1 CD-ROM 12 cm Research report submitted to the U.S. Department of Justice, Award Number: 2009-DN-BX-K180. Ficheiro de 3,15 MB em formato PDF (39 p.) Short tandem repeat (STR) typing at the single-cell level is a promising tool for human forensicidentification when the biological evidence materials are comprised of mixtures of cells from multiple individuals at relatively low concentrations. Here we describe a novel single-cell STR typing method with high sensitivity, fidelity and throughput that combines microfluidic droplet generation with single-cell multiplex emulsion PCR. Individual cells are separately isolated within microdroplets that subsequently function as miniaturized reactors for PCR amplification, producing high quality STR profiles from single cells at high throughput. CONTENTS: 1. Introduction. 2. Methods. 2.1 Microfluidic device fabrication. 2.2 Preparation of primer-functionalized beads. 2.3 Cell culture and sample preparation. 2.4 Microdroplet generation. 2.5 Cell lysis and DNA purification. 2.6 Emulsion PCR. 2.7 Bead recovery. 2.8 Secondary PCR and fragment sizing analysis. 3. Results. 3.1 Microbead-based solid-phase PCR in bulk solutions. 3.2 Agarose microdroplet-based emulsion PCR. 3.3 Single-molecule PCR in microdroplets. 3.4 Single-cell STR typing in microdroplets. 4. Conclusions. 5. References. 6. Dissemination of research findings. IDENTIFICAÇÃO DE PESSOAS 343105 QUÍMICA FORENSE 343048 ANÁLISE LABORATORIAL 326721 GENG Tao 358134 MATHIES Richard A. 358135 Oferta 0 PJCD_258_000000000000000 2022-06-06 PJCD258 426516 PJ PJ PJ1CD258_55 PJCD258 2022-06-06 0 MO Oferta