Contents lists available at ScienceDirect Forensic Science International: Genetics Supplement Series journal homepage: www.elsevier.com/locate/fsigss Preservation of DNA from saliva samples in suboptimal conditions Adria Michelle Burrows, Peter Gustav Ristow, Maria Eugenia D’Amato ⁎ Forensic DNA Laboratory, University of the Western Cape, Cape Town, South Africa ARTICLE INFO Keywords: Preserve Saliva Genomic DNA Storage buffer Population genetics ABSTRACT We designed a storage buffer for preventing DNA from degradation on saliva samples. The efficiency of this buffer to prevent DNA degradation was tested on 10 samples at room temperature for up to 24 months. DNA was extracted using an optimised phenol chloroform method. Agarose gels did not show signs of degradation in DNA after 24 months. Purity ranged from 0.9–1.88 and DNA yield ranged from 2–88 ng/μl. Full profiles were ob- tained from all samples using 1 ng DNA with AmpFlSTR ® Identifiler ® Plus (Applied Biosystems). 1. Introduction Population genetics studies require large sample numbers. When the collection of biological material occurs in remote and isolated locations without facilities, the correct preservation of material is of great con- cern. Saliva has been determined as a reliable, non-invasive source of genomic DNA [1,2]. Several commercial kits are now available for collecting saliva secretions. In this project an economical saliva col- lection method was created to store saliva secretions for extended periods at suboptimal conditions and optimised to compete with ex- isting saliva collection kits. Commercial kits such as Oragene•DIS- COVER (OGR-500) are able to preserve saliva samples up to 5 years in a dark environment at room temperature [3,4]. Our buffer (FDL-storage buffer) was tested in its capacity to preserve saliva at room temperature over several time points between day of collection and 24 months. The integrity and quantity of the extracted DNA samples were evaluated via spectrophotometer analysis, agarose gel electrophoresis, quantification using an intercalating dye qPCR method and genotyping using AmpFlSTR ® Identifiler ® Plus (Applied Biosystems). 2. Material and methods A total of 10 individuals donated saliva samples, which were pre- served with Oragene and our buffer, adding equal volume to approxi- mately 5 ml saliva in a 15 ml polypropylene tube and stored at room temperature. We followed Oragene instructions for collection and DNA extractions. The DNA extractions were conducted using a 500 μl aliquot of the total volume of sample following an optimised phenol chloroform extraction method on the day of collection and then 6, 12 and 24 months afterwards. DNA yield, purity and integrity were evaluated measuring absorbance at 260 nm and 280 nm using a Nanodrop 2000 (Thermo Scientific™), an in-house qPCR method using KAPA HRM kit with intercalating EvaGreen dye, and 0.7% agarose gel electrophoresis respectively. Genotyping with AmpFlSTR ® Identifiler ® Plus (Applied Biosystems) was performed on 1 ng DNA for each sample and run on an ABI 3500 as per the manufacturers protocols and checked using Genemapper, IDX v1.4. 3. Results and discussion The agarose electrophoresis gel in Fig. 1 did not show signs of de- gradation after 24 months. An independent samples t-test was con- ducted to compare the 260/280 purity ratio of the FDL-storage buffer and Oragene storage at all different collection points; t(3) = 3.18, p = 0.73. This result suggests that the purity of the novel FDL-storage buffer is similar to that obtained using Oragene method. Full profiles were obtained after 12 and 24 months of storage (Fig. 2). 4. Conclusion In this work we have shown that the novel FDL-storage is proficient in DNA preservation in suboptimal conditions which will aid future sample collections. Further studies are needed to display the range of conditions at which this buffer can support DNA preservation. Conflict of interest None. http://dx.doi.org/10.1016/j.fsigss.2017.09.050 Received 24 August 2017; Accepted 11 September 2017 ⁎ Corresponding author. E-mail address: medamato@uwc.ac.za (M.E. D’Amato). Forensic Science International: Genetics Supplement Series 6 (2017) e80–e81 Available online 12 September 2017 1875-1768/ © 2017 Elsevier B.V. All rights reserved. T