Aykut Arif TOPÇU, Süleyman AŞIR, Deniz TÜRKMEN

Katı Faz Ayırma Yöntemiyle DNA Saflaştırılması

D NA’nın saflaştırılması veya izole edilmesi moleküler biyoloji, tıbbi çalışmalar ve gen tedavi çalışmaları için önem taşır. Özellikle PCR çalışmaları için yüksek saflıkta DNA molekülüne ihtiyaç duyulmaktadır. Nükleik asitlerin saflaştırılmasında sırasıyla çözücü özütlemesi ve katı faz özütlemesi olmak üzere iki yöntem kulanılır. Çözücü ekstraksiyonu kolay uygulanabilir olmasına rağmen; toksisite, yüksek oranda çözücü ihtiyacı, yöntemin uzun zaman alması ve düşük özgüllüğü gibi birtakım kısıtlamalar mecvuttur. Günümüzde çeşitli örneklerden DNA’nın saflaştırılması için katı faz özütlemesi SPE yaygın olarak kullanılmaktadır. Bu çalışmada; özellikle DNA’nın saflaştırılması için SPE yönteminin uygulamaları üzerinde durulmuş ve bilimsel çalışmalar özetlenmiştir

Anahtar Kelimeler:

DNA, DNA ayrılması, DNA saflaştırılması, SPE

DNA Purification by Solid Phase Extraction SPE Methods

DNA isolation/purification is a widely used procedure for molecular biology, medical studies, gene therapy and especially PCR studies which needs high quality or extremely pure nucleic acid samples. Nucleic acid separation was mainly achieved by two methods; solvent extraction and solid phase extraction SPE methods. Although solvent extraction method is the simplest method, there are some drawbacks such as toxicity, more solvent requirement, time-consuming procedure and low specificity. Nowadays, there is a wide use of solid phase matrices to extract DNA from a variety of samples. In this review article; we basically focused on SPE for DNA purification methods.

Keywords:

DNA, DNA separation, DNA purification, SPE,

PDF

___

D. Cortes-Chosan, R.L. Griffits, Methods for extracting genomic DNA from whole blood samples: current perspectives, J. Biorepository Sci. App. Med, 2 (2014)
B. Rittich, A, Spanova, SPE and purification of DNA using magnetic particles, J. Sep. Sci., 36 (2013) 2472.
N. Ludwig, Nucleic acid techniques in bacterial systematics and identification, Int. J. Food. Microbiol., 120 (2007) 225.
X.Y. Sun, P.Z. Li, B. Ai, Y.B. Wang, Surface modification of MCM-41 and its application in DNA adsorption, Chinese Chem. Lett., 27 (2016) 139.
N.M. Fahrenkopf, P.Z. Rice, M. Bergkvist, N.A. Deskins, and N.C. Cady, Immobilization mechanisms of deoxyribonucleic acid (DNA) to hafnium dioxide (HfO2) surfaces for biosensing applications, Appl. Mater. Interfaces., 4 (2012) 5360.
M.D. Costioli, I. Fisch, F. Flaudy-Garret, F. Hilbrig, R. Freitag, DNA purification by triple-helix affinity precipitation, Biotechnol. Bioeng., 81 (2003) 535.
B. Rittich, A. Spanova, SPE and purification of DNA using magnetic particles, J. Sep. Sci., 36 (2013) 2472.
J. Sambrook, D.W. Russel, Molecular Cloning, 3rd Ed., Cold Spring Harbor Laboratory Press, NY, USA (2001).
M. Wink, An introduction to molecular biotechnology, 2nd Ed, Wiley-VCH Verlag GmbH&Co, KGaA (2011).
M. Deng, C. Jiang, L. Jia, N-Methylimidazolium modified magnetic particles as adsorbents for solid phase extraction of genomic deoxyribonucleic acid from genetically modified soybeans, Anal. Chim. Acta., 771 (2013) 31.
C.F. Poole, New trends in solid-phase extraction, Trends. Anal. Chem., 22 (2003) 362.
E. Yavuz, Ş. Tokalıoğlu, H. Şahan, Ş. Patat, Nanosized spongelike MnO as an adsorbent for 34 preconcentration by vortex assisted solid phase extraction of copper and lead in various food and herb samples, Food Chem., 19 (2016) 463.
N. Bereli, K. Köse, D. Türkmen, A. Denizli, A., Glutamic acid containing supermacroporous poly(hydroxyethyl methacrylate) cryogel disks for uranium removal, Mater. Sci. and Eng. C., 32 (2012) 2052.
C. Armutcu, N. Bereli, E. Bayram, L. Uzun, R. Say, A. Denizli, Aspartic acid incorporated monolithic columns for affinity glycoprotein purification, Colloids and Surfaces B: Biointerfaces., 114, 67-74, 2014.
R. Üzek, L. Uzun, S. Şenel, A. Denizli, Nanospines incorporation into the structure of the hydrophobic cryogels via novel cryogelation method: An alternative sorbent for plasmid DNA purification, Colloid. Surface. B., 102 (2013) 243.
C.L. Chiang, C.S. Sung, C.Y. Chen, Application of silica–magnetite nanocomposites to the isolation of ultrapure plasmid DNA from bacterial cells, J. Magn. Magn. Mater., 305 (2006) 483.
D. H u s k a , Au to m a te d n u c l e i c a c i d s i s o l a ti o n using paramagnetic microparticles coupled with electrochemical detection, Talanta., 79 (2009) 402.
I. Perçin, V. Karakoç, S. Akgöl, E. Aksöz, A. Denizli, Poly(hydroxyethyl methacrylate) based magnetic nanoparticles for plasmid DNA purification from Escherichia coli lysate, Mater. Sci. Eng. C., 32 (2012) 1133.
X.W. Chen, Q.X. Mao, J.W. Liu, J.H. Wang, Isolation/ separationof plasmid DNA using hemoglobin modified magnetic nanocomposites as solid-phase adsorbent, Talanta., 100 (2012) 107.
M.M. Rahman, A. Elaissan, Temperature and magnetic dual responsive microparticles for DNA separation, Sep. Purif. Technol., 81 (2011) 286.
G.M. Whitesides, The origins and the future of microfluidics, Nature., 442 (2006) 368.
E.K. Sackman, A.L. Fulton, & D.J. Beebe, The present and future role of microfluidics in biomedical research, Nature., (2104) 507.
P. Sajeesh, A.K. Sen, Particle separation and sorting in microfluidic devices: a review, Microfluid. Nanofluid., 17 (2014) 1.
A.M. Azimi, et all., A magnetic bead-based DNA extraction and purification microfluidic device, Microfluid. Nanofluid., 11 (2011) 157.
W. Cho, J.H. Maeng, S.Y. Hwang, Disposable on- chip microfluidic system for buccal cell lysis, DNA purification, and polymerase chain reaction, Electrophoresis., 34 (2103) 2531.
Song, H., Wang, Y., Garson, C. Pant, K., Concurrent DNA preconcentration and separation in bipolar electrode-based microfluidic device, Anal. Method., 7 (2015).
A. Denizli, E. Pişkin, Dye-ligand affinity systems, J. Biochem. Biophys. Method., 49 (2001) 391.
M. Wilcheck, My life with affinity, Protein Sci., 13 (2004) 3066.
D. Çimen, F. Yılmaz, I. Perçin, D. Türkmen, A. Denizli, Dye affinity cryogels for plasmid DNA purification, Mater. Sci. and Eng. C., 56 (2015) 318.
F.M. Plieva, I.Y. Galaev, B. Mattiason, B., Macroporous gels prepared at subzero temperatures as novel materials for chromatography of particulate- containing fluids and cell culture applications, J. Sep. Sci., 30 (2007) 1657.
E. Özgür, N. Bereli, D. Türkmen, S. Ünal, A. Denizli, PHEMA cryogel for in-vitro removal of anti-dsDNA antibodies from SLE plasma, Mater. Sci. Eng. C., 31 (2011) 915.
E. Çorman, N. Bereli, S. Özkara, L. Uzun, A. Denizli, Hydrophobic cryogels for DNA adsorption: Effect of embedding of monosize microbeads into cryogel network on their adsorptive performances., Biomed. Chromatogr., 27 (2013) 1524.
J.T. Leonard, M.B. Grace G.S. Buzard, M.J. Mullen, C.B. Barbagallo, Preparation of PCR products for DNA sequencing, Biotechniques., 24 (1998) 314.
R. Westermeier, Electrophoresis in Practice, 4th Ed, Wiley-VCH Verlag GmbH&Co, KGaA, Weinheim (2005).
Tiselius, A., Electrophoresis of serum globulin. Biochem J, 31 (1937). 313.
P. Grabar, C.A. Williams, Method permitting the combined study of the electrophoretic and the immunochemical properties of protein mixtures; application to blood serum, Biochim. Biophys. Acta., 10 (1953) 193.
H.V. Thorne, Electrophoretic separation of polyoma virus DNA from host cell DNA, Virology., 29 (1966) 234.
C. Aaij, P. Borst, The gel electrophoresis of DNA, Biochim Biophys Acta., 269 (1972) 192.
T. Maniatis, A. Jeffrey, H. Van deSande, Chain length determination of small double- and single-stranded DNA molecules by polyacrylamide gel electrophoresis, Biochemistry., 14 (1975) 3787.
A.E. Dahlberg, C.W. Dingman, A.C. Peacock, Electrophoretic characterization of bacterial polyribosomes in agarose-acrylamide composite gels, J Mol. Biol., 41 (1969) 139.
T.J. Kasper, M. Melera, P. Gozel, R.G. Brownlee, Separation and detection of DNA by capillary electrophoresis, J. Chromatogr., 458 (1998) 303.
A.S. Cohen, D. Najarian, J.A. Smith, B.L. Karger, Rapid separation of DNA restriction fragments using capillary electrophoresis, J. Chromatogr., 458 (1988) 323.
D.Y. Chen, K. Adelhelm, X.L. Cheng, N.J. Dovichi, A simple laser-induced fluorescence detector for sulforhodamine 101 in a capillary electrophoresis sysytem: detection limits of 10 yoctomoles or six molecules, Analyst, 11 (1994) 349.
N.J. Dovichi, DNA sequencing by capillary electrophoresis, Electrophoresis, 18 (1997) 2393.
H. Zhang, Z. Wang, X. Li, X.C. Le, Ultrasensitive detection of proteins by amplification of affinity aptamers, Angew. Chem. Int. Ed., 45 (2006) 1576.
E. Fiore, E. Dausse, H. Dubouchaud, E. Peyrin, C. Ravelet, C., Ultrafast capillary electrophoresis isolation of DNA aptamer for the PCR amplification- based small analyte sensing, Front. Chem., 3 (2015) 1.
E.T. Thostenson, Z. Ren, T.S. Chou, Advanced in the science and technology of carbon nanotubes and their composites: a review, Compos. Sci. Technol., 61 (2001) 1899.
P.G. Collins, P. Avouris, Nanotubes for electronics. Sci. Am, 283 (2000) 62.
R.H. Baugman, A.A. Zakhidov, W.A, Heer, W.A., Carbon Nanotubes-the Route Toward Applications, Science, 297 (2002) 787.
I.I. Shakhmaeva, E.R. Bulatov, O.V. Bondar, D.V. Saifullina, M. Culha, A.A. Rizvanov, T.I. Abdullin, Binding and purification of plasmid DNA using multi- layered carbon nanotubes, J. Biotechnol., 152 (2011) 102.
M. Zheng, A. Jagota, E.D. Semke, B.A. Diner, R.S. McLean, R.S., Lustig, S.R., Richardson, R.E., Tassi, N.G., DNA-assisted dispersion and separation of carbon nanotubes, Nature Mater, 2 (2003) 338.
X. Li, Y. Peng, X. Qu, Carbon nanotubes selective destabilization of duplex and triplex DNA and inducing B-A transition in solution, Nucleic Acids Res., 34 (2006) 3670.
F. Nazarian-Firouzabadi, A. Ismaili, S.M. Zabeti, Phenol-stacked carbon nanotubes: A new approach to genomic DNA isolation from plants, Mol. Biol. Res. Commun., 3 (2014) 205.