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Quality of DNA extracted from saliva samples collected with the Oragene™ DNA self-collection kit.

ABSTRACT: Large epidemiological studies in DNA biobanks have increasingly used less invasive methods for obtaining DNA samples, such as saliva collection. Although lower amounts of DNA are obtained as compared with blood collection, this method has been widely used because of its more simple logistics and increased response rate. The present study aimed to verify whether a storage time of 8?months decreases the quality of DNA from collected samples.Saliva samples were collected with an OrageneTM DNA Self-Collection Kit from 4,110 subjects aged 14-15?years. The samples were processed in two aliquots with an 8-month interval between them. Quantitative and qualitative evaluations were carried out in 20% of the samples by spectrophotometry and genotyping. Descriptive analyses and paired t-tests were performed.The mean volume of saliva collected was 2.2?mL per subject, yielding on average 184.8??g DNA per kit. Most samples showed a Ratio of OD differences (RAT) between 1.6 and 1.8 in the qualitative evaluation. The evaluation of DNA quality by TaqMan®, High Resolution Melting (HRM), and restriction fragment length polymorphism-PCR (RFLP-PCR) showed a rate of success of up to 98% of the samples. The sample store time did not reduce either the quantity or quality of DNA extracted with the Oragene kit.The study results showed that a storage period of 8?months at room temperature did not reduce the quality of the DNA obtained. In addition, the use of the Oragene kit during fieldwork in large population-based studies allows for DNA of high quantity and high quality.

SUBMITTER: Nunes AP

PROVIDER: S-EPMC3422993 | biostudies-literature | 2012 May

REPOSITORIES: biostudies-literature

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Quality of DNA extracted from saliva samples collected with the Oragene™ DNA self-collection kit.

Nunes Ana P AP Oliveira Isabel O IO Santos Betânia R BR Millech Cristini C Silva Liziane P LP González David A DA Hallal Pedro C PC Menezes Ana M B AM Araújo Cora L CL Barros Fernando C FC

BMC medical research methodology 20120504

<h4>Background</h4>Large epidemiological studies in DNA biobanks have increasingly used less invasive methods for obtaining DNA samples, such as saliva collection. Although lower amounts of DNA are obtained as compared with blood collection, this method has been widely used because of its more simple logistics and increased response rate. The present study aimed to verify whether a storage time of 8 months decreases the quality of DNA from collected samples.<h4>Methods</h4>Saliva samples were co ...[more]

PMID: 22559914

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Project description:BackgroundTechnical 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.MethodsSaliva 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.ResultsThe smallest tested whole saliva volume of .10 ml yielded, on average, 1.43 ± .77 μg 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.ConclusionsWhole 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.

Dataset Information

Quality of DNA extracted from saliva samples collected with the Oragene™ DNA self-collection kit.

Publications

Quality of DNA extracted from saliva samples collected with the Oragene™ DNA self-collection kit.

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