Solid-phase extraction nucleic acid

The biological applications of NMR include the study of the structure of macromolecules such as proteins and nucleic acids and the study of membranes, and enzymic reactions. Newer methods and instruments have overcome, to a large extent, the technical difficulties encountered with aqueous samples and the analysis of body fluids is possible, permitting the determination of both the content and concentration of many metabolites in urine and plasma. NMR is not a very sensitive technique and it is often necessary to concentrate the sample either by freeze drying and dissolving in a smaller volume cm- by solid phase extraction methods. 

Wolfe, K.A., Breadmore, M.C., Ferrance, J.P., Power, M.E., Conroy, J.F., Norris, P.M., Landers, J.P., Toward a microchip-based solid-phase extraction method for isolation of nucleic acids. Electrophoresis 2001, 23, 727-733. 

One application example on the microfluidic LSI platform is the extraction of nucleic acids (NA) from a small amount of cells [126, 127] for cell-based assays. For the extraction of NA from a cell suspension, the cell membrane has to be destroyed first (chemical lysis of the cell). Afterwards, the NA are specifically separated from the residual cell components using a solid phase extraction method based on an NA affinity column (paramagnetic beads). This extraction protocol is completely implemented on the microfiuidic... 

Chitosans find a wide variety of applications in chromatographic separations. The presence of free amino and hydroxyl groups in chitosan makes it a useful chromatographic support. Chitosan can be used in thin layer chromatography for separation of nucleic acid and solid phase extraction of phenols and chlorophenols [69]. 

Reedy CR, Hagan KA, Strachan BC, Higginson JJ, Bienvenue JM, Greenspoon S A, Ferrance JP, Landers JP (2010) Dual-domain microchip-based process for volume reduction solid phase extraction of nucleic acids from dilute, large volume biological samples. Anal Chem 82 5669-5678... 

T.-H. Wang, Y. Zhang, Fabrication of hierarchical silica nanomembranes and uses thereof for solid phase extraction of nucleic acids, US 2015/0,037,802 Al, 2015. 

Quake and coworkers [16] developed a PDMS microfluidic device (shown in Eig. 4c) for nucleic acid purification from a small number of bacterial or mammalian cells. This multilayer device contained fluidic channels and a system of membrane-actuated pneumatic valves and pumps, which enabled precise control of buffers, lysis agents and cell solution, and also allowed for parallel processing. Bacterial cells, dilution buffer and lysis buffer are first introduced into the chip and then transferred into the rotary mixer. Once mixed, the lysate is flushed over a DNA affinity column and drained. The DNA is recovered from the chip with an elution buffer for further analysis. We note that this is the only microfluidic chemical C3flome-try device to use a separation method other than solution-phase electrophoresis (i. e., solid phase extraction). 

Fast blot methods to minimize nucleic acid extraction and immobilization steps have been developed. Those with nylon as a solid phase can take advantage of the ability of NaOH to dissociate cells, denature DNA and immobilize DNA. Nitrocellulose membranes have a lower binding capacity and co-immobilization of nucleic acid and protein from neutral solutions can be a problem. Bresser et al. (1983) used hot concentrated Nal to inhibit protein immobilization, to denature DNA and to irreversibly bind the nucleic acid to nitrocellulose (no baking required). This method can also be used for RNA. About 10 cells are minimally required for a unique DNA sequence, whereas > 0.01% of total mRNA can be detected by the Nal methods. 

Several kits are commercially available for the extraction of RNA from plant, bacteria, or mammalian sources. We have used the RNeasy Mini Kit (Qiagen, Valencia, CA) for the extraction of RNA from bacteria following the procedure recommended by the manufacturer (92). This kit relies on guanidine thiocyanate-silica isolation of RNA. Here, guanidine thiocyanate serves as a chaotropic agent, which both lyses cells and inactivates nucleases. In its presence, the released nucleic acids bind to silica particles, which provide a solid phase from which the collected RNA can later be eluted using water (93). This kit is suitable for the isolation and purification of up to 100 pg RNA molecules >200... 

A variety of well-established macroscale SPE methods for nucleic acid extraction have been successfully transferred to microscale devices [10, 31-57]. Although the physical principles of these methods may be different (e.g., chaotropic interactions, electrostatic interactions, affinity interactions, etc.), micro-SPE protocols typically consist of three steps (1) selective adsorption of nucleic acids onto a solid phase (2) removal of contaminants by a washing step and (3) elution of the preconcentrated nucleic acids from the solid support using water or a low salt buffer [31]. Like their macroscale counterparts, micro-SPE devices possess a loading level of target material that is dependent upon the available surface area within the extraction bed and, thus, are manufactured either by packing the solid phase... 

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