Rapid multiplex DNA amplification on an inexpensive microdevice for human identification via short tandem repeat analysis

DuVall, Jacquelyn A.; Roux, Delphine Le; Thompson, Brandon L.; Birch, Christopher; Nelson, Daniel A.; Li, Jingyi; Mills, Daniel L.; Tsuei, An‐Chi; Ensenberger, Martin G.; Sprecher, Cindy; Storts, Douglas R.; Root, Brian E.; Landers, James P.

doi:10.1016/j.aca.2017.04.051

Cited by 16 publications

(9 citation statements)

References 27 publications

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“…Lower total volumes have been shown to help reduce the time of heating and cooling of the sample, reducing amplification time by 56–73% . DuVall et al used a 10 loci multiplex containing a subset of the CODIS loci (all smaller than 350 BP) to achieve amplification in 15 min . This microfluidic chip could be coupled with conventional or nonconventional extraction and detection methods to achieve the users’ requirements.…”

Section: Genotyping Methods Using Short Tandem Repeatsmentioning

confidence: 99%

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Forensic DNA Analysis

McCord

Gauthier

Cho³

et al. 2018

Anal. Chem.

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Section: Genotyping Methods Using Short Tandem Repeatsmentioning

confidence: 99%

“…75 DuVall et al used a 10 loci multiplex containing a subset of the CODIS loci (all smaller than 350 BP) to achieve amplification in 15 min. 76 This microfluidic chip could be coupled with conventional or nonconventional extraction and detection methods to achieve the users' requirements.…”

Section: ■ Dna Extraction and Sample Recoverymentioning

confidence: 99%

Forensic DNA Analysis

McCord

Gauthier

Cho³

et al. 2018

Anal. Chem.

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“…Several prominent studies have even developed and tested microdevices for processing sexual assault samples [12][13][14][15][16]. The goal of maintaining simplicity with these microdevices usually manifests in the implementation of a direct cell lysis assay, rather than conventional DNA extraction and purification [17][18][19]. Regardless of the approach, processing of these samples always requires the addition of DTT or another reducing agent to sufficiently lyse any sperm cells present.…”

mentioning

confidence: 99%

The effects of dithiothreitol (DTT) on fluorescent qPCR dyes

Hudson

Cox

Seashols‐Williams

et al. 2020

Journal of Forensic Sciences

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DNA extractions of semen samples commonly utilize dithiothreitol (DTT) to reduce and disrupt disulfide bonds. Although traditional extraction techniques remove DTT before downstream analyses, the forensic DNA community has recently explored Y‐screening, direct amplification, and direct cell lysis assays that omit purification but employ reducing agents to lyse spermatozoa. This study examined the impact of residual DTT on downstream processes involving fluorescent dyes. Quantification using Investigator® Quantiplex HYres revealed a significant increase in the male DNA yield (p = 0.00056) and a >150,000,000‐fold increase in the male:human DNA ratio when DTT remained in extracts versus when it was filtered out using a traditional purification method. When DTT was present with Quantifiler™ Trio, the true mean DNA yield for the large autosomal target significantly increased (p = 0.038) and the average reported DNA yields increased 1.1‐fold, >9.5‐fold, and 1.3‐fold for the small autosomal, large autosomal, and male targets, respectively. DTT‐spiked DNA standards from both kits were impacted similarly to samples with residual DTT, demonstrating that observed effects were related to DTT and not the extraction method. This study corroborates other reports that DTT adversely affects multiple dyes (e.g., Cy5, Quasar 670, SYBR Green I, TMR, and Mustang Purple®). Overall, DTT causes inaccurate quantities and, consequently, inaccurate calculated male:female ratios when used in conjunction with these kits. Thus, implementation of newer direct‐to‐PCR assays incorporating DTT should either be avoided or used only with carefully evaluated, compatible dyes.

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“…Samples were prepared following a previously described protocol [12]. Briefly, DNA from a buccal swab containing saliva was enzymatically extracted using Micro-GEM reagents (MicroGEM International, PLC, VA, USA).…”

Section: Sample Extraction and Amplificationmentioning

confidence: 99%

Rapid, inexpensive fabrication of electrophoretic microdevices for fluorescence detection

et al. 2022

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The laser print, cut, and laminate (PCL) method for microfluidic device fabrication can be leveraged for rapid and inexpensive prototyping of electrophoretic microchips useful for optimizing separation conditions. The rapid prototyping capability allows the evaluation of fluidic architecture, applied fields, reagent concentrations, and sieving matrix, all within the context of using fluorescencecompatible substrates. Cyclic olefin copolymer and toner-coated polyethylene terephthalate (tPeT) were utilized with the PCL technique and bonding methods optimized to improve device durability during electrophoresis. A series of separation channel designs and centrifugation conditions that provided successful loading of sieving polymer in less than 3 min was described. Separation of a 400-base DNA sizing ladder provided calculated base resolution between 3 and 4 bases, a greater than 18-fold improvement over separations on similar substrates. Finally, the accuracy and precision capabilities of these devices were demonstrated by separating and sizing DNA fragments of 147 and 167 bases as 148.62 ± 2 and 166.48 ± 3 bases, respectively.

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Rapid multiplex DNA amplification on an inexpensive microdevice for human identification via short tandem repeat analysis

Cited by 16 publications

References 27 publications

Forensic DNA Analysis

Forensic DNA Analysis

The effects of dithiothreitol (DTT) on fluorescent qPCR dyes

Rapid, inexpensive fabrication of electrophoretic microdevices for fluorescence detection

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