Search results for "DNA extraction"
showing 10 items of 66 documents
Molecular characterization of dominant bacterial population in “Vastedda della Valle del Belice” cheese: preliminary investigation
2010
The sensory characteristics of raw-milk cheeses are linked to the cheese-making process, to the environmental factors as animal feeding and to the biochemical and microbiological composition of the milk. In this report we temped to characterize the microflora in the typical Sicilian historical cheese as Vastedda della valle del Belice. Each cheese was previous subjected to microbial isolation on specific media (M17 and MRS). The colony obtained on the solid medium were subject to biochemical tests and DNA extraction. The microbial diversity occurring in the strains was evaluated by PCR, RFLP and sequencing targeted on 16S ribosomal DNA. A number of closest relatives species of lactic and co…
A METHOD FOR STUDYING GENETICS OF THE PREDATORY MITE PHYTOSEIULUS PERSIMILIS
2008
Two DNA extraction and purification procedures for single mite analysis, organic (phenol-chloroform) method and the Chelex® -100 DNA extraction procedure, have been compared concerning extraction efficiency, DNA purity and DNA suitability for amplification. Phenol-chloroform extraction takes a relatively long period of time (5 days) and involves the use of toxic and hazardous chemical reagents but it produces consistent results. Chelex is a chelating resin that has a high affinity for polyvalent metal ions, Singer-Sam et al. postulated that boiling a sample in the presence of chelex prevents the degradation of DNA. Both method were evaluated to extract DNA from single frozen mite or individ…
Presence and characterisation of verotoxin producing E. coli in fresh Italian pork sausages, and preparation and use of an antibiotic-resistant strai…
2005
One hundred and twenty six samples of fresh pork sausages were analysed-for the presence of verocytotoxigenic Escherichia coli (VTEC). Selective enrichment followed by DNA extraction and PCR amplification of the stx1 and stx2 genes highlighted the occurrence of the above mentioned genes in 20 out of 126 samples screened. From the stx positive enriched cultures, isolation was performed on CT-SMAC agar plates after immuno-magnetic separation of E. coli O157. Fifty three non-sorbitol fermenting isolates were obtained and further characterised, along with the reference strain E. coli ATCC 35150(T). All the isolates were characterised by PCR assays, assessing the presence of stx1, stx2, rfbE(O15…
Application of an inexpensive and high-throughput genomic DNA extraction method for the molecular ecology of zooplanktonic diapausing eggs
2008
We describe the application of a simple, low-cost, and effective method of DNA extraction (hot sodium hydroxide and Tris, HotSHOT) to the diapausing propagules of continental aquatic invertebrates for its use in PCR amplification. We illustrate the use of the technique in cladocerans, rotifers, anostracans, notostracans, and copepod diapausing eggs. We compare the performance of the HotSHOT technique to the currently most widely used method for DNA extraction of zooplankton eggs and individuals, the chelating resin (or Chelex) technique. The HotSHOT technique overcomes several of the problems posed by Chelex and permits easy optimization for its use with 96-well plates for high-throughput D…
Inter-laboratory evaluation of the ISO standard 11063 "Soil quality -- Method to directly extract DNA from soil samples"
2011
International audience; Extracting DNA directly from micro-organisms living in soil is a crucial step for the molecular analysis of soil microbial communities. However, the use of a plethora of different soil DNA extraction protocols, each with its own bias, makes accurate data comparison difficult. To overcome this problem, a method for soil DNA extraction was proposed to the International Organization for Standardization (ISO) in 2006. This method was evaluated by 13 independent European laboratories actively participating in national and international ring tests. The reproducibility of the standardized method for molecular analyses was evaluated by comparing the amount of DNA extracted, …
Biotechnological applications of the sepiolite interactions with bacteria: Bacterial transformation and DNA extraction
2020
International audience; Among the various clay minerals, sepiolite, which is a natural nanofibrous silicate that exhibit a poor cell toxicity, is a potential promising nanocarrier for the non-viral and stable transfer of plasmid DNA into bacteria, mammalian and human cells. We first show here that sepiolite binds to bacteria, which can be useful in decontamination protocols. In a previous research we have shown that is possible to modulate the efficiency of the absorption of different types of DNA molecules onto sepiolite, and that the DNA previously adsorbed could be recovered preserving the DNA structure and biological activity. Taking advantage of both, the sepiolite/bacteria and sepioli…
Biogeographical patterns of soil molecular microbial biomass
2011
Affiche - résumé. Session GC: Microbes in the Changing Environmenl: Global Climate Change and Soif under Human Impact; International audience; The spatial organization of soil microbial communities over large areas and the identification of environmental factors structuring their distribution have been liUle investigated. The overall objective of this study was to determine the spatial paUerning of microbial biomass in soils on wide extent and to rank the environmental fiUers most influencing this distribution, by using the French Soil Quality Monitoring Network. This network covers ail the French terrilory and soils were sampled at2,150 sites along a systematic grid sampling. The soil DNA …
DNA Extraction From Orthoptera Museum Specimens
2011
We describe a procedure for rapid purification of high quality DNA from either fresh or dry Orthoptera, suitable for the PCR amplification of DNA regions more than 800 bp long (even from oldest specimens), which allows genetic analyses on animals from collections without the complete specimen disruption.
Determination of steady-state levels of oxidative DNA base modifications in mammalian cells by means of repair endonucleases
1997
The alkaline elution technique in combination with various repair endonucleases (Fpg protein, endonuclease III, exonuclease III, T4 endonuclease V) was used to quantify steady-state (background) levels of oxidative base modifications in various types of mammalian cells. In human lymphocytes the number of base modifications sensitive to Fpg protein, which include 8-hydroxyguanine, was 0.25 +/- 0.05 per 10(6) base pairs. Even lower levels (0.07 +/- 0.02 per 10(6) bp) were observed in HeLa cells. The numbers of sites sensitive to the other repair endonucleases were below the detection limit (0.05 per 10(6) bp). In a direct comparison, the background level of Fpg-sensitive modifications determi…
Electrophoretic pattern of the polypeptide component in the tight DNA-protein complexes in rat and chicken tissues and its partial characterisation i…
2010
Electrophoretic pattern of the polypeptide component in the tight DNA-protein complexes in rat and chicken tissues and its partial characterisation in rat liverTightly bound to DNA proteins (TBP) are a protein group that remain attached to DNA with covalent or non-covalent bonds after its deproteinisation. The distribution of TBP in genes reflects the type of cell differentiation. It has been hypothesised that TBP binding is involved in regulation of gene expression. Early studies reported uniformity of the TBPs from different sources. Later it was shown that TBPs obtained from DNA, isolated in mild conditions from evolutionary distant species, are different. Application of chloroform DNA e…