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Mol Cell Probes. 2011 Feb;25(1):44-8. doi: 10.1016/j.mcp.2010.10.003. Epub 2010 Oct 26.

Genomic DNA extraction from whole blood stored from 15- to 30-years at -20 °C by rapid phenol-chloroform protocol: a useful tool for genetic epidemiology studies.

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1
School of Biosciences and Biotechnologies, Via Gentile III da Varano, University of Camerino, 62032 Camerino, Italy.

Abstract

Long-term stored (LTS) whole blood collection can be an important source of DNA without collection costs, but there is a lack of information on methods useful to extract genomic DNA from such type of biological material. Here we report a simple and fast revisited phenol/chloroform extraction method from LTS whole blood. Protocol reliability was assessed by comparison with proteinase K and silica-gel membrane spin column-based DNA extraction methods using LTS -20 °C whole blood from 1980, and by testing it on 82 whole blood samples, collected from 1980 to 1995, with high quality (A(260/280) = 1.79 ± 0.32 O.D., A(260/230) = 1.45 ± 0.52 O.D.) and quantity results. Genotyping efficiency was also checked by performing RFLP-PCR and ASP-PCR of p53 Pro72Arg (rs1042522) SNP and hTERT MNS16A VNTR, respectively, resulting in 100% of samples successfully typed. In addition to the goodness and the efficiency of method proposed here, this protocol achieves working time reduction combining extraction and purification steps, allowing to work at room temperature. Furthermore, phenol is able to inactivate any potential nuclease and potential infective sources from the first step on. Based on these results we also conclude that LTS -20 °C whole blood samples may be considered a reliable and potential resource for future genotyping studies and retrospective analysis in a genetic epidemiological setting.

PMID:
21029772
DOI:
10.1016/j.mcp.2010.10.003
[Indexed for MEDLINE]

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