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The utility of DNA extracted from saliva for genome-wide molecular research platforms.

ABSTRACT: The study aimed to investigate the suitability of DNA extracted from saliva for high throughput molecular genotyping and DNA methylation platforms by comparing its performance with that of DNA extracted from blood. The genome-wide methylation profile, using the Infinium HumanMethylation450 Beadchip array® (Illumina, San Diego, CA), was measured for 20 DNA samples. Common genetic variation was measured, using the Infinium HumanCore Beadchip® (Illumina, San Diego, CA) for 4 samples (matching samples from 2 people).DNA from blood and saliva returned genotyping call rates and reproducibility frequencies of > 99%. High-quality DNA methylation data was obtained from both saliva and blood DNA, with average detection p-values for each sample ranging from 0.001 to 0.006. Slightly higher global DNA methylation levels were observed in whole blood DNA than saliva DNA. Correlations between individuals for each sample type were generally greater than correlations between two sample types from the same individual (Pearson's correlation, r = 0.9696 in 10 pairs of matched blood and saliva derived DNA, r = 0.9702 between saliva samples, and r = 0.9769 between blood derived DNA). Saliva yields DNA of sufficient quantity and quality to compare favourably with blood as a source of DNA for genetic and epigenetic research purposes.

SUBMITTER: Bruinsma FJ

PROVIDER: S-EPMC5759806 | biostudies-other | 2018 Jan

REPOSITORIES: biostudies-other

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The utility of DNA extracted from saliva for genome-wide molecular research platforms.

Bruinsma Fiona J FJ Joo Jihoon E JE Wong Ee Ming EM Giles Graham G GG Southey Melissa C MC

BMC research notes 20180108 1

<h4>Objective</h4>The study aimed to investigate the suitability of DNA extracted from saliva for high throughput molecular genotyping and DNA methylation platforms by comparing its performance with that of DNA extracted from blood. The genome-wide methylation profile, using the Infinium HumanMethylation450 Beadchip array<sup>®</sup> (Illumina, San Diego, CA), was measured for 20 DNA samples. Common genetic variation was measured, using the Infinium HumanCore Beadchip<sup>®</sup> (Illumina, San ...[more]

PMID: 29310721

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Project description:BACKGROUND: Technical advances following the Human Genome Project revealed that high-quality and -quantity DNA may be obtained from whole saliva samples. However, usability of previously collected samples and the effects of environmental conditions on the samples during collection have not been assessed in detail. In five studies we document the effects of sample volume, handling and storage conditions, type of collection device, and oral sampling location, on quantity, quality, and genetic assessment of DNA extracted from cells present in saliva. METHODS: Saliva samples were collected from ten adults in each study. Saliva volumes from .10-1.0 ml, different saliva collection devices, sampling locations in the mouth, room temperature storage, and multiple freeze-thaw cycles were tested. One representative single nucleotide polymorphism (SNP) in the catechol-0-methyltransferase gene (COMT rs4680) and one representative variable number of tandem repeats (VNTR) in the serotonin transporter gene (5-HTTLPR: serotonin transporter linked polymorphic region) were selected for genetic analyses. RESULTS: The smallest tested whole saliva volume of .10 ml yielded, on average, 1.43 ± .77 ?g DNA and gave accurate genotype calls in both genetic analyses. The usage of collection devices reduced the amount of DNA extracted from the saliva filtrates compared to the whole saliva sample, as 54-92% of the DNA was retained on the device. An "adhered cell" extraction enabled recovery of this DNA and provided good quality and quantity DNA. The DNA from both the saliva filtrates and the adhered cell recovery provided accurate genotype calls. The effects of storage at room temperature (up to 5 days), repeated freeze-thaw cycles (up to 6 cycles), and oral sampling location on DNA extraction and on genetic analysis from saliva were negligible. CONCLUSIONS: Whole saliva samples with volumes of at least .10 ml were sufficient to extract good quality and quantity DNA. Using 10 ng of DNA per genotyping reaction, the obtained samples can be used for more than one hundred candidate gene assays. When saliva is collected with an absorbent device, most of the nucleic acid content remains in the device, therefore it is advisable to collect the device separately for later genetic analyses.

Project description:DNA methylation has become increasingly recognized in the etiology of psychiatric disorders. Because brain tissue is not accessible in living humans, epigenetic studies are most often conducted in blood. Saliva is often collected for genotyping studies but is rarely used to examine DNA methylation because the proportion of epithelial cells and leukocytes varies extensively between individuals. The goal of this study was to evaluate whether saliva DNA is informative for studies of psychiatric disorders. Saliva and blood methylation was clearly distinguishable though there was positive correlation overall. There was little correlation in CpG sites within relevant candidate genes. Correlated CpG sites were more likely to occur in areas of low CpG density (i.e. CpG shores and open seas). There was more variability in CpG sites from saliva than blood, which may reflect its heterogeneity. Thus, this study provides a framework for using DNA methylation from saliva and suggests that DNA methylation of saliva may offer distinct opportunities for epidemiological and longitudinal studies of psychiatric traits. DNA methylation was assessed in saliva and blood samples from 64 adult African Americans. Both saliva and blood samples were collected from each participant. Saliva was stored in Oragene DNA sample collection kits (DNA Genotek), and blood was collected in EDTA vacuum tubes. DNA was extracted using the Puregene Genomic DNA kit (Invitrogen). DNA methylation was interrogated for each sample using the HumanMethylation450 BeadChip (Illumina). Analyses for tissue-specific DNA patterns were conducted using linear regression adjusted for appropriate covariates, including estimated cellular proportions. The estimated proportion of epithelial cells in saliva DNA ranged from 3-99% (median 26%). In blood, the estimated proportion of lymphocytes ranged from 25-70% (median 48%) and neutrophils ranged from 42-84% (median 58%). Methylation of 68.8% of all CpG sites differed relative to epithelial cell proportion (FDR<.05). Our results provide a framework for methylation studies in DNA extracted from saliva and highlight the importance of controlling for the proportion of epithelial and leukocyte cells. This presents an attractive opportunity for investigators that have already collected salivary DNA for genetic studies of psychiatric traits.

Dataset Information

The utility of DNA extracted from saliva for genome-wide molecular research platforms.

Publications

The utility of DNA extracted from saliva for genome-wide molecular research platforms.

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