Nucleic extraction

Synthesized SAMNPs, in conjunction with basic chemistry for standard nucleic extraction and purification, can be used to capture both the crude cells during prelysis (to concentrate the targeted cells) and subsequently capture genomic DNA for postlysis (to purify... 

Extraction of proteia requires breaking the cell wall to release the cytoplasmic contents. This can be achieved by high speed ball or coUoid mills or by high pressure (50—60 Mpa) extmsion. Proteia is extracted by alkaline treatment followed by precipitation after enzymatic hydrolysis of nucleic acids. Although the proteia can be spun iato fibers or texturized, such products are more expensive than those derived from soybean and there is no market for them. 

The presence of nucleic acids ia yeast is oae of the maia problems with their use ia human foods. Other animals metabolize uric acid to aHantoia, which is excreted ia the uriae. Purines iagested by humans and some other primates are metabolized to uric acid, which may precipitate out ia tissue to cause gout (37). The daily human diet should contain no more than about 2 g of nucleic acid, which limits yeast iatake to a maximum of 20 g. Thus, the use of higher concentrations of yeast proteia ia human food requires removal of the nucleic acids. Unfortunately, yields of proteia from extracts treated as described are low, and the cost of the proteia may more than double. 

Lipoteichoic acids (from gram-positive bacteria) [56411-57-5J. Extracted by hot phenol/water from disrupted cells. Nucleic acids that were also extracted were removed by treatment with nucleases. Nucleic resistant acids, proteins, polysaccharides and teichoic acids were separated from lipoteichoic acids by anion-exchange chromatography on DEAE-Sephacel or by hydrophobic interaction on octyl-Sepharose [Fischer et al. Ear J Biochem 133 523 1983]. 

The previous biomarkers relate to phenotypic assessments of microbial diversity and most will probably measure a restricted part of the total microbial pool, since not alt markers will be expressed uniformly by every cell. In contrast, methods involving the detection of nucleic acids may be directly applicable to all microorganisms provided that the complete extraction of DNA (lysis of cells) or permea-bilization of cells can be achieved. 

Classical approaches to plant DNA isolation aim to produce large quantities of highly purified DNA. However, smaller quantities of crudely extracted plant DNA are often acceptable for PCR analysis. Another efficient method for preparation of plant DNA for PCR is a single-step protocol that involves heating a small amount of plant tissue in a simple solution. Several factors influence nucleic acid release from tissue salt, EDTA, pH, incubation time and temperature. These factors must be optimized for different sample substrates. EDTA in the sample solution binds the Mg + cofactor required by the Taq polymerase in the PCR, so the EDTA concentration in the solution, or the Mg + concentration in the PCR, must be carefully optimized. 

Seasonal variations in the metabolic fate of adenine nucleotides prelabelled with [8—1-4C] adenine were examined in leaf disks prepared at 1-month intervals, over the course of 1 year, from the shoots of tea plants (Camellia sinensis L. cv. Yabukita) which were growing under natural field conditions by Fujimori et al.33 Incorporation of radioactivity into nucleic acids and catabolites of purine nucleotides was found throughout the experimental period, but incorporation into theobromine and caffeine was found only in the young leaves harvested from April to June. Methy-lation of xanthosine, 7-methylxanthine, and theobromine was catalyzed by gel-filtered leaf extracts from young shoots (April to June), but the reactions could not be detected in extracts from leaves in which no synthesis of caffeine was observed in vivo. By contrast, the activity of 5-phosphoribosyl-1-pyrophosphate synthetase was still found in leaves harvested in July and August. 

Nucleic acid extraction protocols using guanidine hydrochloride, sodium sarco-syl, and ethanol have been developed to quantify viral RNA by bDNA in lymph node tissue, liver tissue, and peripheral blood monocytes (Wilber and Urdea, 1995). 

Burtscher, C. Fall, P. A. Wilderer, P. A. Wuertz, S. Detection of Salmonella and Listeria monocytogenes in suspended organic waste by nucleic acid extraction and PCR. Appl. Environ. Microbiol. 1999, 65, 2235-2237. 

Sample preparation used to extract proteins from cells prior to analysis is an important step that can have an effect on the accuracy and reproducibility of the results. Proteins isolated from bacterial cells will have co-extracted contaminants such as lipids, polysaccharides, and nucleic acids. In addition various organic salts, buffers, detergents, surfactants, and preservatives may have been added to aid in protein extraction or to retain enzymatic or biological activity of the proteins. The presence of these extraneous materials can significantly impede or affect the reproducibility of analysis if they are not removed prior to analysis. 

This chapter will address the applications of protein-based bioinformatics to analysis of microorganisms introduced intact into the instrumental system for rapid processing and analysis. Strategies that require offline extraction and fractionation of proteins will not be discussed. Although the amplification of nucleic acids is a powerful approach, especially coupled with mass spectrometry,15 it requires extraction and processing, and thus is not included. 

Subsequently, similar experiments were done with viral nucleic acids. The pure viral nucleic acid, when added to cells, led to the synthesis of complete virus particles the protein coat was not required. This process is called transfection. More recently, DNA has been used in cell-free extracts to program the synthesis of RNA that functions as the template for the synthesis of proteins characteristic of the DNA... 

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