title: Improvement and automation of a real-time PCR assay for vaginal fluids.
abstract: The identification of vaginal fluids is crucial in forensic science. Several molecular protocols based on PCR amplification of mfDNA (microflora DNA) specific for vaginal bacteria are now available. Unfortunately mfDNA extraction and PCR reactions require manual optimization of several steps. The aim of present study was the verification of a partial automatization of vaginal fluids identification through two instruments widely diffused in forensic laboratories: EZ1 Advanced robot and Rotor Gene Q 5Plex HRM. Moreover, taking advantage of 5-plex thermocycler technology, the ForFluid kit performances were improved by expanding the mfDNA characterization panel with a new bacterial target for vaginal fluids and with an internal positive control (IPC) to monitor PCR inhibition. Results underlined the feasibility of a semi-automated extraction of mfDNA using a BioRobot and demonstrated the analytical improvements of the kit.
authors.affiliation: University of Rome "Foro Italico", Department of Movement, Human and Health Sciences, Public Health Unit, Piazza Lauro De Bosis, 6, 00135 Rome, Italy; Legal Genetics - forensic consulting, Via Po, 102, 00198 Rome, Italy.
authors.forename: E
authors.initials: E
authors.lastname: De Vittori
authors.affiliation: University of Rome "Foro Italico", Department of Movement, Human and Health Sciences, Public Health Unit, Piazza Lauro De Bosis, 6, 00135 Rome, Italy.
authors.forename: S
authors.initials: S
authors.lastname: Giampaoli
authors.affiliation: Carabinieri, Reparto Investigazioni Scientifiche di Roma, Viale di Tor di Quinto, 119, 00191 Rome, Italy.
authors.forename: F
authors.initials: F
authors.lastname: Barni
authors.affiliation: Legal Genetics - forensic consulting, Via Po, 102, 00198 Rome, Italy.
authors.forename: M
authors.initials: M
authors.lastname: Baldi
authors.affiliation: Carabinieri, Reparto Investigazioni Scientifiche di Roma, Viale di Tor di Quinto, 119, 00191 Rome, Italy.
authors.forename: A
authors.initials: A
authors.lastname: Berti
authors.affiliation: Carabinieri, Reparto Investigazioni Scientifiche di Roma, Viale di Tor di Quinto, 119, 00191 Rome, Italy.
authors.forename: L
authors.initials: L
authors.lastname: Ripani
authors.affiliation: University of Rome "Foro Italico", Department of Movement, Human and Health Sciences, Public Health Unit, Piazza Lauro De Bosis, 6, 00135 Rome, Italy. Electronic address: vincenzo.romanospica@uniroma4.it.
authors.forename: V
authors.initials: V
authors.lastname: Romano Spica
mesh_terms: D000328:Adult; D000368:Aged; D001331:Automation; D004269:DNA, Bacterial / Q000235:genetics* / Q000302:isolation & purification; D005240:Feasibility Studies; D005260:Female; D005554:Forensic Medicine; D006801:Humans; D008875:Middle Aged; D060888:Real-Time Polymerase Chain Reaction; D012463:Saliva / Q000382:microbiology; D014621:Vagina / Q000382:microbiology*
chemical_list: D004269:DNA, Bacterial
keywords: Automated DNA extraction; Multiplex PCR; Vaginal fluid; mfDNA