A novel, integrated forensic microdevice on a rotation-driven platform: Buccal swab to STR product in less than 2 h.
This work describes the development of a novel microdevice for forensic DNA processing of reference swabs. This microdevice incorporates an enzyme-based assay for DNA preparation, which allows for faster processing times and reduced sample handling. Infrared-mediated PCR (IR-PCR) is used for STR amplification using a custom reaction mixture, allowing for amplification of STR loci in 45 min while circumventing the limitations of traditional block thermocyclers. Uniquely positioned valves coupled with a simple rotational platform are used to exert fluidic control, eliminating the need for bulky external equipment. All microdevices were fabricated using laser ablation and thermal bonding of PMMA layers. Using this microdevice, the enzyme-mediated DNA liberation module produced DNA yields similar to or higher than those produced using the traditional (tube-based) protocol. Initial microdevice IR-PCR experiments to test the amplification module and reaction (using Phusion Flash/SpeedSTAR) generated near-full profiles that suffered from interlocus peak imbalance and poor adenylation (significant -A). However, subsequent attempts using KAPA 2G and Pfu Ultra polymerases generated full STR profiles with improved interlocus balance and the expected adenylated product. A fully integrated run designed to test microfluidic control successfully generated CE-ready STR amplicons in less than 2 h (<1 h of hands-on time). Using this approach, high-quality STR profiles were developed that were consistent with those produced using conventional DNA purification and STR amplification methods. This method is a smaller, more elegant solution than current microdevice methods and offers a cheaper, hands-free, closed-system alternative to traditional forensic methods.
Department of Forensic Science, Virginia Commonwealth University, Richmond, VA, USA.
Jordan O
JO
Cox
Department of Forensic Science, Virginia Commonwealth University, Richmond, VA, USA.
Teresa Sikes
TS
DeCarmen
Departments of Chemistry, Mechanical and Aerospace Engineering, & Pathology, University of Virginia, Charlottesville, VA, USA.
Yiwen
Y
Ouyang
Departments of Chemistry, Mechanical and Aerospace Engineering, & Pathology, University of Virginia, Charlottesville, VA, USA.
Briony
B
Strachan
Departments of Chemistry, Mechanical and Aerospace Engineering, & Pathology, University of Virginia, Charlottesville, VA, USA.
Hillary
H
Sloane
Department of Forensic Science, Virginia Commonwealth University, Richmond, VA, USA.
Cathey
C
Connon
Department of Forensic Science, Virginia Commonwealth University, Richmond, VA, USA.
Kemper
K
Gibson
Departments of Chemistry, Mechanical and Aerospace Engineering, & Pathology, University of Virginia, Charlottesville, VA, USA.
Kimberly
K
Jackson
Departments of Chemistry, Mechanical and Aerospace Engineering, & Pathology, University of Virginia, Charlottesville, VA, USA.
James P
JP
Landers
Department of Forensic Science, Virginia Commonwealth University, Richmond, VA, USA.
Tracey Dawson
TD
Cruz
D004247:DNA / Q000032:analysis* / Q000235:genetics; D004867:Equipment Design; D053803:Forensic Genetics / Q000379:methods*; D006801:Humans; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation* / Q000379:methods*; D018895:Microsatellite Repeats / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods*
D004247:DNA
Fast PCR; Forensic DNA; Microdevice; Microfluidics; Rapid DNA