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Extraction Kits (extraction + kit)
Selected AbstractsAn optimized method to separate reticulocytes from peripheral blood for molecular analysisINTERNATIONAL JOURNAL OF LABORATORY HEMATOLOGY, Issue 3 2009R. PETRUZZELLI Summary A method based on immunomagnetic sorting of reticulocytes from peripheral blood was set up and combined to a commercial extraction kit for the isolation of total RNA from whole blood. This procedure resulted in high-quality RNA samples suitable for molecular analysis. We used this procedure to analyse erythroid-specific transcripts, starting from peripheral blood samples, to search for differently expressed mRNAs in patients with hereditary persistence of foetal haemoglobin. After erythrocyte lysis, CD15+and CD45+ peripheral cells were negatively sorted to remove leucocyte populations that could have affected the subsequent screening procedure. The cell sorting and RNA extraction procedure was completed within 1,2 h of erythrocyte lysis, which represents a consistent saving of time compared with other procedures. Moreover, it produced 1 ,g of total RNA per ml of blood samples, which is sufficient for molecular analysis. Therefore, our method is a reliable and efficient tool to isolate RNA from specific cell subpopulations poorly represented in peripheral blood, particularly when accurate detection and characterization of highly unstable and poorly expressed molecules is required. [source] DNA Extraction from Olive Oil and PCR Amplification of Microsatellite MarkersJOURNAL OF FOOD SCIENCE, Issue 1 2005Raffaele Testolin And ABSTRACT: DNA was extracted from single-cultivar of cold-pressed (virgin) unfiltered and cotton-filtered olive oils that were stored at 4 °C for up to a year using different DNA extraction kits and protocols. DNA was amplified using original and nested primers designed on 6 microsatellites loci of the UDO series. The most consistent results in terms of successful single sequence repeat amplifications were achieved using the Qiagen QIAamp DNA stool extraction kit, slightly modified and applied to oil sample amounts as small as 200 ,L without any pretreatment. The kit allowed getting polymerase chain reaction (PCR) amplicons visible on gel and scorable peaks at the automatic sequencer for all 6 markers analyzed. Less consistent results were achieved with other kits, such as the Promega Wizard Magnetic DNA Purification System for Food, the LB Link-Biotech ExtMan 50,100 Evolution, the Qiagen Plant Mini kit, and the standard cetyltrimethyl-ammonium bromide-based DNA extraction protocol. The integration in the protocols of further tools, such as the hexane-based phase separation, the addition of water or NaCl solutions to the oil, the precipitation and the use of the pellet, and others, did not result in any substantial use. PCR amplifications that gave low DNA yields were improved by adopting the nested PCR technique, which uses the product of the 1st PCR as a template for a 2nd PCR carried out by means of internal primers. Conclusions are drawn as to the applicability of the method to trace the identity of single-cultivar virgin olive oils. Further work is required to check the sensitivity of the method in determining the varietal composition of blended oils, especially in detecting alleles from cultivars present in only small amounts. [source] Evaluation and optimisation of five different extraction methods for soy DNA in chocolate and biscuits.JOURNAL OF THE SCIENCE OF FOOD AND AGRICULTURE, Issue 11 2004Extraction of DNA as a first step in GMO analysis Abstract A method is described to discriminate between genetically modified (GM) and non-modified foodstuffs by detecting the presence of newly introduced genes at the protein or DNA level. Currently available methods operate almost exclusively at the DNA level and are based on the polymerase chain reaction (PCR). The first and most crucial step in this process is the isolation of DNA. In this study, five different methods for the isolation of DNA from chocolate and biscuits were evaluated, using four commercially available extraction kits and a non-commercial method for amplification of the soybean-specific lectin gene. The latter method involves the use of hot-start Taq polymerase, to prevent the formation of non-specific amplification products, and an increase in the number of cycles from 35 to 41. The performance of the non-commercial cetyl trimethylammonium bromide (CTAB)-based method was the best, taking into consideration the adaptations of the extraction procedure, although this method was more time-consuming than the others. Chocolate (white, milk and dark) and several biscuits generated positive amplification results using this PCR approach. Copyright © 2004 Society of Chemical Industry [source] Recovery of plant DNA using a reciprocating saw and silica-based columnsMOLECULAR ECOLOGY RESOURCES, Issue 1 2007PATRICK J. ALEXANDER Abstract The time needed for hand grinding and the cost of commercially available extraction kits remain to be the major limitations in plant DNA extraction for many researchers. We present inexpensive techniques for (i) simultaneously machine grinding large numbers of plant samples for DNA extraction using a commercially available reciprocating saw; and (ii) DNA recovery using silica column-based extractions similar to that used in some commercially available kits. Used together, these allow for the rapid recovery of plant DNA at relatively low cost. Furthermore, these methods appear to be widely applicable within plants with good yields recovered in test extractions across major plant groups (ferns, gymnosperms, monocots and eudicots). [source] | ||||||||||