Molecular microbiological methods, such as competetive PCR, real-time PCR, denaturing gradient gel electrophoresis (DGGE) and large-scale parallel-pyrosequencing, require the extraction of sufficient quantity of high ...Molecular microbiological methods, such as competetive PCR, real-time PCR, denaturing gradient gel electrophoresis (DGGE) and large-scale parallel-pyrosequencing, require the extraction of sufficient quantity of high quality DNA from microbiologically and chemically complex matrices. Due to difficulties in the field to standardize/select the optimum DNA preservation-extraction methods in view of laboratories differences, this article attempts to present a straight-forward mathematical framework for comparing some of the most commonly used methods. To this end, as a case study, the problem of selecting an optimum sample preservation-DNA extraction strategy for obtaining total bacterial DNA from swine feces was considered. Two sample preservation methods (liquid nitrogen and RNAlater?) and seven extraction techniques were paired and compared under six quantitative DNA analysis criteria: yield of extraction, purity of extracted DNA (A260/280 and A 260/230 ratios), duration of extraction, degradation degree of DNA, and cost. From a practical point of view, it is unlikely that a single sample preservation-DNA extraction strategy can be optimum for all selected criteria. Hence, a systematic multi-criteria decision-making (MCDM) approach was used to compare the methods. As a result, the ZR Fecal DNA MiniPrepTM DNA extraction kit for samples preserved either with liquid nitrogen or RNAlater? were identified as potential optimum solutions for obtaining total bacterial DNA from swine feces. Considering the need for practicality for in situ applications, we would recommend liquid nitrogen as sample preservation method, along with the ZR Fecal DNA MiniPrepTM kit. Total bacterial DNA obtained by this strategy can be suitable for downstream PCR-based DNA analyses of swine feces.展开更多
There is an increased interest in the extraction of nucleic acids from various environmental samples since culture-independent molecular techniques contribute to deepen and broaden the understanding of a greater porti...There is an increased interest in the extraction of nucleic acids from various environmental samples since culture-independent molecular techniques contribute to deepen and broaden the understanding of a greater portion of uncultivable microorganisms. Due to difficulties to select the optimum DNA extraction method in view of downstream molecular analyses, this article presents a straightforward mathematical framework for comparing some of the most commonly used methods. Four commercial DNA extraction kits and two physical-chemical methods (bead-beating and freeze-thaw) were compared for the extraction of DNA under several quantitative DNA analysis criteria: yield of extraction, purity of extracted DNA (A260/280 and A260/230 ratios), degradation degree of DNA, easiness of PCR amplification, duration of extraction, and cost per extraction. From a practical point of view, it is unlikely that a single DNA extraction strategy can be optimum for all selected criteria. Hence, a systematic Technique for Order Preference by Similarity to Ideal Solution (TOPSIS) was employed to compare the methods. The PowerSoil? DNA Isolation Kit was systematically defined as the best performing method for extracting DNA from soil samples. More specifically, for soil:manure and soil:manure:biochar mixtures, the PowerSoil?DNA Isolation Kit method performed best, while for neat soil samples its alternative version gained the first rank.展开更多
Recognition of the biological properties of numerous “natural products” has fueled the current focus of this field, namely, the search for new drugs, antibiotics, insecticides, and herbicides. Based on their biosynt...Recognition of the biological properties of numerous “natural products” has fueled the current focus of this field, namely, the search for new drugs, antibiotics, insecticides, and herbicides. Based on their biosynthetic origins, natural products can be divided into three major groups: the isoprenoids, alkaloids, and phenolic compounds. Isoprenoids are structurally the most diverse group of secondary natural metabolites with different roles in the growth, development, and reproduction of a diverse range of prokaryotic and eukaryotes cells. Mevalonate and 2-C-methyl-D-erythritol 4-phosphate (MEP) pathways are known to be responsible for biosynthesis of numerous isoprenoids. HMG-CoA reductase is a rate-determining enzyme in mevalonate pathway, producing intermediates such as farnesyl and geranylgeranyl pyrophosphates, which lead to by-products such as cholesterol. Earlier studies have demonstrated that the inhibition of HMG-CoA reductase is one of the most effective approaches for treating hypercholesterolemia and eventually cardiovascular disease (CVD). Statins are HMG-CoA reductase inhibitors and the most prescribed group of drugs worldwide in treating hypercholesterolemia;however the application of this group of drugs may be expensive and has side effects including rashes and gastrointestinal symptoms. For these reasons, there is an important need to examine the viability of natural products as an alternative to statin treatment. This article is a review of different aforementioned areas with a focus on isoprenoids that can be used for the regulation of HMG-CoA reductase.展开更多
文摘Molecular microbiological methods, such as competetive PCR, real-time PCR, denaturing gradient gel electrophoresis (DGGE) and large-scale parallel-pyrosequencing, require the extraction of sufficient quantity of high quality DNA from microbiologically and chemically complex matrices. Due to difficulties in the field to standardize/select the optimum DNA preservation-extraction methods in view of laboratories differences, this article attempts to present a straight-forward mathematical framework for comparing some of the most commonly used methods. To this end, as a case study, the problem of selecting an optimum sample preservation-DNA extraction strategy for obtaining total bacterial DNA from swine feces was considered. Two sample preservation methods (liquid nitrogen and RNAlater?) and seven extraction techniques were paired and compared under six quantitative DNA analysis criteria: yield of extraction, purity of extracted DNA (A260/280 and A 260/230 ratios), duration of extraction, degradation degree of DNA, and cost. From a practical point of view, it is unlikely that a single sample preservation-DNA extraction strategy can be optimum for all selected criteria. Hence, a systematic multi-criteria decision-making (MCDM) approach was used to compare the methods. As a result, the ZR Fecal DNA MiniPrepTM DNA extraction kit for samples preserved either with liquid nitrogen or RNAlater? were identified as potential optimum solutions for obtaining total bacterial DNA from swine feces. Considering the need for practicality for in situ applications, we would recommend liquid nitrogen as sample preservation method, along with the ZR Fecal DNA MiniPrepTM kit. Total bacterial DNA obtained by this strategy can be suitable for downstream PCR-based DNA analyses of swine feces.
文摘There is an increased interest in the extraction of nucleic acids from various environmental samples since culture-independent molecular techniques contribute to deepen and broaden the understanding of a greater portion of uncultivable microorganisms. Due to difficulties to select the optimum DNA extraction method in view of downstream molecular analyses, this article presents a straightforward mathematical framework for comparing some of the most commonly used methods. Four commercial DNA extraction kits and two physical-chemical methods (bead-beating and freeze-thaw) were compared for the extraction of DNA under several quantitative DNA analysis criteria: yield of extraction, purity of extracted DNA (A260/280 and A260/230 ratios), degradation degree of DNA, easiness of PCR amplification, duration of extraction, and cost per extraction. From a practical point of view, it is unlikely that a single DNA extraction strategy can be optimum for all selected criteria. Hence, a systematic Technique for Order Preference by Similarity to Ideal Solution (TOPSIS) was employed to compare the methods. The PowerSoil? DNA Isolation Kit was systematically defined as the best performing method for extracting DNA from soil samples. More specifically, for soil:manure and soil:manure:biochar mixtures, the PowerSoil?DNA Isolation Kit method performed best, while for neat soil samples its alternative version gained the first rank.
文摘Recognition of the biological properties of numerous “natural products” has fueled the current focus of this field, namely, the search for new drugs, antibiotics, insecticides, and herbicides. Based on their biosynthetic origins, natural products can be divided into three major groups: the isoprenoids, alkaloids, and phenolic compounds. Isoprenoids are structurally the most diverse group of secondary natural metabolites with different roles in the growth, development, and reproduction of a diverse range of prokaryotic and eukaryotes cells. Mevalonate and 2-C-methyl-D-erythritol 4-phosphate (MEP) pathways are known to be responsible for biosynthesis of numerous isoprenoids. HMG-CoA reductase is a rate-determining enzyme in mevalonate pathway, producing intermediates such as farnesyl and geranylgeranyl pyrophosphates, which lead to by-products such as cholesterol. Earlier studies have demonstrated that the inhibition of HMG-CoA reductase is one of the most effective approaches for treating hypercholesterolemia and eventually cardiovascular disease (CVD). Statins are HMG-CoA reductase inhibitors and the most prescribed group of drugs worldwide in treating hypercholesterolemia;however the application of this group of drugs may be expensive and has side effects including rashes and gastrointestinal symptoms. For these reasons, there is an important need to examine the viability of natural products as an alternative to statin treatment. This article is a review of different aforementioned areas with a focus on isoprenoids that can be used for the regulation of HMG-CoA reductase.
