Rapid extraction and preservation of genomic DNA from human samples

AbstractSimple and rapid extraction of human genomic DNA remains a bottleneck for genome analysis and disease diagnosis. Current methods using microfilters require cumbersome, multiple handling steps in part because salt conditions must be controlled for attraction and elution of DNA in porous silica. We report a novel extraction method of human genomic DNA from buccal swab and saliva samples. DNA is attracted onto a gold-coated microchip by an electric field and capillary action while the captured DNA is eluted by thermal heating at 70 °C. A prototype device was designed to handle four microchips, and a compatible protocol was developed. The extracted DNA using microchips was characterized by qPCR for different sample volumes, using different lengths of PCR amplicon, and nuclear and mitochondrial genes. In comparison with a commercial kit, an equivalent yield of DNA extraction was achieved with fewer steps. Room-temperature preservation for 1 month was demonstrated for captured DNA, facilitating straightforward collection, delivery, and handling of genomic DNA in an environment-friendly protocol. FigurePortable microtip device for human genomic DNA extraction

[1]  J. Wagner,et al.  Characterization of monocrotaline pyrrole-induced DNA cross-linking in pulmonary artery endothelium. , 1993, The American journal of physiology.

[2]  T. Lindahl,et al.  Rate of depurination of native deoxyribonucleic acid. , 1972, Biochemistry.

[3]  Bruce Budowle,et al.  Assessing a novel room temperature DNA storage medium for forensic biological samples. , 2012, Forensic science international. Genetics.

[4]  G. Csako.  Present and future of rapid and/or high-throughput methods for nucleic acid testing. , 2006, Clinica chimica acta; international journal of clinical chemistry.

[5]  T. Lindahl,et al.  Heat-induced depyrimidination of deoxyribonucleic acid in neutral solution. , 1973, Biochemistry.

[6]  M. Bissell.  DNA yield and quality of saliva samples and suitability for large-scale epidemiological studies in children , 2012 .

[7]  Woon-Hong Yeo,et al.  Electric field-induced concentration and capture of DNA onto microtips , 2012 .

[8]  C. Frontali,et al.  The alkaline denaturation of DNA. , 1969, Biophysical journal.

[9]  Steven A. Soper,et al.  Purification and preconcentration of genomic DNA from whole cell lysates using photoactivated polycarbonate (PPC) microfluidic chips , 2006, Nucleic acids research.

[10]  William A. Mcmillan,et al.  Rapid, automated nucleic acid probe assays using silicon microstructures for nucleic acid concentration. , 1999, Journal of biomechanical engineering.

[11]  A. Fornace,et al.  Analysis of the effect of DNA alkylation on alkaline elution. , 1986, Carcinogenesis.

[12]  R. Shapiro,et al.  The deamination of cytidine and cytosine by acidic buffer solutions. Mutagenic implications. , 1966, Biochemistry.

[13]  M. Otte,et al.  Isolation of genomic DNA from buccal swabs for forensic analysis, using fully automated silica-membrane purification technology. , 2003, Legal medicine.

[14]  H. Morgan,et al.  Dielectrophoresis of DNA: time- and frequency-dependent collections on microelectrodes , 2006, IEEE Transactions on NanoBioscience.

[15]  Marco Rito-Palomares,et al.  Dielectrophoresis for the manipulation of nanobioparticles , 2007, Electrophoresis.

[16]  M. Smits,et al.  Automated genomic DNA extraction from saliva using the QIAxtractor , 2010, Clinical Chemistry and Laboratory Medicine.

[17]  R. Pethig.  Review article-dielectrophoresis: status of the theory, technology, and applications. , 2010, Biomicrofluidics.

[18]  T. Anchordoquy,et al.  Preservation of DNA , 2007 .

[19]  K. A. Wolfe,et al.  Toward a microchip‐based solid‐phase extraction method for isolation of nucleic acids , 2002, Electrophoresis.

[20]  Christophe Frippiat,et al.  Evaluation of novel forensic DNA storage methodologies. , 2011, Forensic science international. Genetics.

[21]  J. Bonnet,et al.  Chain and conformation stability of solid-state DNA: implications for room temperature storage , 2009, Nucleic acids research.

[22]  Woon-Hong Yeo,et al.  Immunosensor towards low-cost, rapid diagnosis of tuberculosis. , 2012, Lab on a chip.

[23]  Daniel C Leslie,et al.  Nucleic Acid Extraction Techniques and Application to the Microchip , 2009 .

[24]  M J Carter,et al.  An inexpensive and simple method for DNA purifications on silica particles. , 1993, Nucleic acids research.

[25]  K. Buetow,et al.  Collection of genomic DNA from adults in epidemiological studies by buccal cytobrush and mouthwash. , 2001, Cancer epidemiology, biomarkers & prevention : a publication of the American Association for Cancer Research, cosponsored by the American Society of Preventive Oncology.