Evaluation of different sources of DNA for use in genome wide studies and forensic application

Safar, Habiba S. Al; Abidi, Fatima H.; Khazanehdari, Kamal; Dadour, Ian R.; Tay, Guan K.

doi:10.1007/s00253-010-2926-3

Cited by 12 publications

(12 citation statements)

References 22 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…A small amount of serum (250 µL) can be very informative, since it yields enough DNA to analyse several genetic markers. Note that plasma or serum is more reliable than whole blood for clinical studies, specifically for retrospective studies based on stored DNA and for shipment of sera in clinical trial analysis with centralised laboratories [18]. The present study also evidenced the possibility of using DNA from serum and plasma to simulate serum or plasma utilization for DNA analysis of stored samples.…”

Section: Discussionmentioning

confidence: 54%

“…Sample collection does not require highly trained personnel or an established ‘cold chain’ to preserve samples until DNA is extracted. FTA™ cards are a viable storage matrix from DNA, and can be extracted to perform Genome Wide Analysis Studies analysis (GWAS) [18]. Moreover, GWAS are commonly used to identify genetic predispositions of many human diseases.…”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Detection of IL28B SNP DNA from Buccal Epithelial Cells, Small Amounts of Serum, and Dried Blood Spots

et al. 2012

View full text Add to dashboard Cite

Background & AimsPoint mutations in the coding region of the interleukin 28 gene (rs12979860) have recently been identified for predicting the outcome of treatment of hepatitis C virus infection. This polymorphism detection was based on whole blood DNA extraction. Alternatively, DNA for genetic diagnosis has been derived from buccal epithelial cells (BEC), dried blood spots (DBS), and genomic DNA from serum. The aim of the study was to investigate the reliability and accuracy of alternative routes of testing for single nucleotide polymorphism allele rs12979860CC.MethodsBlood, plasma, and sera samples from 200 patients were extracted (400 µL). Buccal smears were tested using an FTA card. To simulate postal delay, we tested the influence of storage at ambient temperature on the different sources of DNA at five time points (baseline, 48 h, 6 days, 9 days, and 12 days)ResultsThere was 100% concordance between blood, plasma, sera, and BEC, validating the use of DNA extracted from BEC collected on cytology brushes for genetic testing. Genetic variations in HPTR1 gene were detected using smear technique in blood smear (3620 copies) as well as in buccal smears (5870 copies). These results are similar to those for whole blood diluted at 1/10. A minimum of 0.04 µL, 4 µL, and 40 µL was necessary to obtain exploitable results respectively for whole blood, sera, and plasma. No significant variation between each time point was observed for the different sources of DNA. IL28B SNPs analysis at these different time points showed the same results using the four sources of DNA.ConclusionWe demonstrated that genomic DNA extraction from buccal cells, small amounts of serum, and dried blood spots is an alternative to DNA extracted from peripheral blood cells and is helpful in retrospective and prospective studies for multiple genetic markers, specifically in hard-to-reach individuals.

show abstract

Section: Discussionmentioning

confidence: 54%

Section: Discussionmentioning

confidence: 99%

Detection of IL28B SNP DNA from Buccal Epithelial Cells, Small Amounts of Serum, and Dried Blood Spots

et al. 2012

View full text Add to dashboard Cite

show abstract

“…R is the normalized intensity value and log R ratio is log to the observed R divided by the expected R [18]. The log R ratio can be used as a measure of noise in the genotyping data [19]. Each genotyped SNP has a corresponding log R ratio.…”

Section: Resultsmentioning

confidence: 99%

“…However, the amplification process is potentially non-uniform, causing regions to be miss-represented. At the same time array data from amplified samples are far more noisy compared to array data from unamplified DNA [14], [15], [19]. To evaluate the noise introduced by the WGA procedure in our samples, we compared genotype array data from genomic DNA samples and their corresponding WGA DNA samples.…”

Section: Resultsmentioning

confidence: 99%

Evaluation of whole genome amplified DNA to decrease material expenditure and increase quality

Bækvad-Hansen

Bybjerg‐Grauholm

Poulsen

et al. 2017

Molecular Genetics and Metabolism Reports

View full text Add to dashboard Cite

AimThe overall aim of this study is to evaluate whole genome amplification of DNA extracted from dried blood spot samples. We wish to explore ways of optimizing the amplification process, while decreasing the amount of input material and inherently the cost. Our primary focus of optimization is on the amount of input material, the amplification reaction volume, the number of replicates and amplification time and temperature. Increasing the quality of the amplified DNA and the subsequent results of array genotyping is a secondary aim of this project.MethodsThis study is based on DNA extracted from dried blood spot samples. The extracted DNA was subsequently whole genome amplified using the REPLIg kit and genotyped on the PsychArray BeadChip (assessing > 570,000 SNPs genome wide). We used Genome Studio to evaluate the quality of the genotype data by call rates and log R ratios.ResultsThe whole genome amplification process is robust and does not vary between replicates. Altering amplification time, temperature or number of replicates did not affect our results. We found that spot size i.e. amount of input material could be reduced without compromising the quality of the array genotyping data. We also showed that whole genome amplification reaction volumes can be reduced by a factor of 4, without compromising the DNA quality.DiscussionWhole genome amplified DNA samples from dried blood spots is well suited for array genotyping and produces robust and reliable genotype data. However, the amplification process introduces additional noise to the data, making detection of structural variants such as copy number variants difficult. With this study, we explore ways of optimizing the amplification protocol in order to reduce noise and increase data quality. We found, that the amplification process was very robust, and that changes in amplification time or temperature did not alter the genotyping calls or quality of the array data. Adding additional replicates of each sample also lead to insignificant changes in the array data. Thus, the amount of noise introduced by the amplification process was consistent regardless of changes made to the amplification protocol. We also explored ways of decreasing material expenditure by reducing the spot size or the amplification reaction volume. The reduction did not affect the quality of the genotyping data.

show abstract

“…A study in 2011 [22] compared three different sources of DNA for GWAS using forensic human samples (degraded genomic DNA (both amplified and unamplified) and amplified DNA from FTA Whatman cards). This study also showed that DNA extracted from FTA cards with subsequent amplification was suitable for GWA studies however, STR PCR was not performed to confirm that no allele drop out or amplification bias had occurred.…”

Section: Discussionmentioning

confidence: 99%

Novel approach for deriving genome wide SNP analysis data from archived blood spots

et al. 2012

View full text Add to dashboard Cite

BackgroundThe ability to transport and store DNA at room temperature in low volumes has the advantage of optimising cost, time and storage space. Blood spots on adapted filter papers are popular for this, with FTA (Flinders Technology Associates) Whatman™TM technology being one of the most recent. Plant material, plasmids, viral particles, bacteria and animal blood have been stored and transported successfully using this technology, however the method of porcine DNA extraction from FTA Whatman™TM cards is a relatively new approach, allowing nucleic acids to be ready for downstream applications such as PCR, whole genome amplification, sequencing and subsequent application to single nucleotide polymorphism microarrays has hitherto been under-explored.FindingsDNA was extracted from FTA Whatman™TM cards (following adaptations of the manufacturer’s instructions), whole genome amplified and subsequently analysed to validate the integrity of the DNA for downstream SNP analysis. DNA was successfully extracted from 288/288 samples and amplified by WGA. Allele dropout post WGA, was observed in less than 2% of samples and there was no clear evidence of amplification bias nor contamination. Acceptable call rates on porcine SNP chips were also achieved using DNA extracted and amplified in this way.ConclusionsDNA extracted from FTA Whatman cards is of a high enough quality and quantity following whole genomic amplification to perform meaningful SNP chip studies.

show abstract

Evaluation of different sources of DNA for use in genome wide studies and forensic application

Cited by 12 publications

References 22 publications

Detection of IL28B SNP DNA from Buccal Epithelial Cells, Small Amounts of Serum, and Dried Blood Spots

Detection of IL28B SNP DNA from Buccal Epithelial Cells, Small Amounts of Serum, and Dried Blood Spots

Evaluation of whole genome amplified DNA to decrease material expenditure and increase quality

Novel approach for deriving genome wide SNP analysis data from archived blood spots

Contact Info

Product

Resources

About