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Saliva as an alternative source of high yield canine genomic DNA for genotyping studies.

ABSTRACT: BACKGROUND: The domestic dog presents an attractive model system for the study of the genetic basis of disease. The development of resources such as the canine genome sequence and SNP genotyping platforms has allowed for the implementation of canine genetic studies. Successful implementation of such studies depends not only on the quality of individual DNA samples, but also on the number of samples obtained. The latter can be maximized using a non-invasive DNA collection method that can increase study participation. We compared the DNA yield and quality obtained from blood and buccal swabs to those obtained using a non-invasive saliva collection kit (Oragene *ANIMAL kit). We also assessed the success rate of PCR amplification and genotyping accuracy of DNA isolated using these collection methods. FINDINGS: Comparison of DNA yields from matched saliva, blood and buccal swab samples showed that yields from saliva were significantly higher than those from blood (p = 0.0198) or buccal swabs (p = 0.0008). Electrophoretic analysis revealed that blood and saliva produced higher quality DNA than buccal swabs. In addition, a 1.1-kb PCR fragment was successfully amplified using the paired DNA samples and genotyping by PCR-RFLP yielded identical results. CONCLUSION: We demonstrate that DNA yields from canine saliva are higher than those from blood or buccal swabs. The quality of DNA extracted from saliva is sufficient for successful amplification of a 1.1-kb fragment and for accurate SNP genotyping by PCR-RFLP. We conclude that saliva presents a non-invasive alternative source of high quantities of canine genomic DNA suitable for genotyping studies.

SUBMITTER: Mitsouras K

PROVIDER: S-EPMC2781016 | biostudies-other | 2009

REPOSITORIES: biostudies-other

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Saliva as an alternative source of high yield canine genomic DNA for genotyping studies.

Mitsouras Katherine K Faulhaber Erica A EA

BMC research notes 20091029

<h4>Background</h4>The domestic dog presents an attractive model system for the study of the genetic basis of disease. The development of resources such as the canine genome sequence and SNP genotyping platforms has allowed for the implementation of canine genetic studies. Successful implementation of such studies depends not only on the quality of individual DNA samples, but also on the number of samples obtained. The latter can be maximized using a non-invasive DNA collection method that can i ...[more]

PMID: 19874604

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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

Saliva as an alternative source of high yield canine genomic DNA for genotyping studies.

Publications

Saliva as an alternative source of high yield canine genomic DNA for genotyping studies.

Similar Datasets

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