title: Optimization of Diamond Nucleic Acid Dye for quantitative PCR.
abstract: Here, we evaluate Diamond Nucleic Acid Dye (DD) for use in quantitative PCR (qPCR) applications. Although DD is a commercially available stain for detection of DNA separated by gel electrophoresis, its use as a detection dye in qPCR has yet to be described. To determine if DD can be used in qPCR, we investigated its inhibitory effects on qPCR at concentrations ranging 0.1-2.5×. Serial dilution of DNA was used to determine the efficiency, sensitivity, and linearity of DD-generated qPCR data in comparison to other commonly used fluorescent dyes such as SYBR Green (SG), EvaGreen (EG), and BRYT Green (BG). DD was found to be comparable with other dyes for qPCR applications, with an R2 value >0.9 and an efficiency of 0.83. Mitochondrial DNA (mtDNA) target signals were successfully produced by DD over a DNA dilution range of ~28 ng- 0.28 pg, demonstrating comparable sensitivity to the other dyes investigated. Cq values obtained using DD were lower than those using EG by almost 7 cycles. We conclude that Diamond Nucleic Acid Dye is a cheaper, less toxic alternative for qPCR applications.
authors.affiliation: School of Biological Sciences, Flinders University, Adelaide, Australia.
authors.forename: Alicia M
authors.initials: AM
authors.lastname: Haines
authors.affiliation: School of Biological Sciences, Flinders University, Adelaide, Australia.|Department of Chemistry and Physics, Arcadia University, Glenside, PA.
authors.forename: Shanan S
authors.initials: SS
authors.lastname: Tobe
authors.affiliation: School of Biological Sciences, Flinders University, Adelaide, Australia.
authors.forename: Adrian
authors.initials: A
authors.lastname: Linacre
mesh_terms: D004247:DNA / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D005456:Fluorescent Dyes / Q000032:analysis* / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D016014:Linear Models; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
chemical_list: D005456:Fluorescent Dyes; D004247:DNA
keywords: Diamond nucleic acid dye; Real-time PCR; SYBR Green; quantitative PCR (qPCR)