In molecular biology, high-quality, purified nucleic acids are the key to nucleic acid analysis. Suitable extraction and purification methods that meet the needs of downstream applications such as blotting, PCR, cloning and sequencing are vital for successful completion of experiments. The steps of nucleic acid extraction include disruption of cells to produce lysates, removal of membrane lipids, proteins, and other nucleic acids, nucleic acid purification/binding from bulk, and concentration of nucleic acids. Cell disruption can be achieved through physical methods, chemical methods, and a combination of the two, with the main aim of destroying the cell wall and/or the cell membrane. The selection of an appropriate destruction method is mainly based on the properties of the sample. The main components of chemical lysis are lytic enzymes, chaotropic agents, and different detergents, while mechanical method disrupts cells by grinding, shearing, bead beating, and shocking.
Methods for DNA and RNA extraction can be broadly characterized into solution-based and solid-phase methods. Solution-based methods rely on the biochemical properties of cellular components to enable the desired molecules to be separated, and may have a preference or exclusivity in extracting DNA or RNA. These methods include cesium chloride (CsCl) gradient centrifugation with ethidium bromide (EtBr), guanidinium thiocyanate- (GuSCN-) phenol-chloroform extraction, cetyltrimethylammonium bromide nucleic acid (CTAB) extraction, chelate resin-based extraction, and alkaline extraction. Both the GuSCN-phenol-chloroform extraction and the CsCl gradient centrifugation with EtBr methods can be used to produce high purity and yield of DNA or RNA, but the former uses hazardous chemicals and the latter is laborious, time-consuming, and costly. Compared to the conventional liquid-phase extraction, the solid-phase extraction is simple, fast, high-throughput, and requires small sample. Solid-phase extraction is a highly efficient nucleic acid extraction technique, which is based on liquid and stationary phases. It selectively separates the target analyte based on specific hydrophobic, polar, and/or ionic properties of the solute and sorbent. Solid-phase extraction methods can be divided into three types: normal/conventional, reverse and ion-exchange. Commonly used solid supports for solid phase extraction include silica matrices, glass particles, diatomaceous earth, anion-exchange carriers, and magnetic beads.
The choice of purification mothed depends on the volume and number of samples, the required turnaround time, the status of nucleic acid fragments, and the organizational and multidimensional formats of nucleic acids (such as genomic, plasmid, tRNA, mRNA, and rRNA). Amerigo Scientific offers a wide range of DNA and RNA extraction and purification products to help you spend less time purifying nucleic acids and more time developing experiments and analyzing data.
.
