Search results for "Gel electrophoresi"
showing 10 items of 321 documents
Genetic diversity and phenotypic characterization of Iodobacter limnosediminis associated with skin lesions in freshwater fish
2021
The relatively unknown genus Iodobacter sp. has been repeatedly isolated from skin ulcers and saprolegniosis on freshwater fish in Finland, especially farmed salmonids. Genetic characterization verified that all 23 bacterial isolates studied here belonged to the species Iodobacter limnosediminis, previously undescribed from the fish microbiota. Whole-genome pulsed-field gel electrophoresis revealed variability between the I. limnosediminis strains, suggesting that they were most likely of environmental origin. Two I. limnosediminis strains caused lesions in 27%–53% of brown trout (Salmo trutta) injected intramuscularly (p ≤ .05). The lesions represented moderate to severe tissue damage, but…
Separation of deoxyribonucleases (DNases) of normal human stratum corneum and psoriatic scales by micro-disc-electrophoresis.
1975
Normal stratum corneum and psoriatic scales were homogenized and a differential centrifugation was performed. The DNase activity of the individual fractions was investigated by micro-disc-electrophoresis. At pH 5 only in the 600 × g pellet and 105.000 × g supernatant of normal keratin DNase activity could be observed. However, all psoriatic fractions showed distinct enzyme activities. At pH 7.4 little psoriatic DNase activity could only be demonstrated in the 105.000 × g supernatant. Except from the 15.000 × g pellet all fractions of normal stratum corneum displayed marked activities. In addition the 105.000 × g supernatant showed two different DNase bands.
Identification of picobirnavirus from faeces of Italian children suffering from acute diarrhea
1996
Polyacrylamide gel electrophoresis of nucleic acid extracted from stool samples of diarrhoeic children revealed in 3 out of 690 (0.43 %) specimens two electrophoretic bands with a migration pattern characteristic of picobirnavirus ds-RNA. In none of the 92 control children were similar bands detected. No other potential enteric pathogens were found in the patients with picobirnavirus infection.
Expression and purification of polyhistidine-tagged rotavirus NSP4 proteins in insect cells
2003
The rotavirus nonstructural NSP4 protein, a transmembrane endoplasmic reticulum-specific glycoprotein, has been described as the first viral enterotoxin. Purified NSP4 or a peptide corresponding to NSP4 residues 114-135 induces diarrhea in young mice. NSP4 has a membrane-destabilizing activity and causes an increase in intracellular calcium levels and chloride secretion by a calcium-dependent signalling pathway in eucaryotic cells. In this study, four recombinant baculoviruses were generated expressing the rotavirus NSP4 glycoprotein from the human strains Wa and Ito, the porcine strain OSU, and the simian strain SA11, which belong to two different NSP4 genotypes, A and B. The recombinant g…
Purification of Large Cytosolic Proteases for In Vitro Assays: 20S and 26S Proteasomes
2012
Proteasomes are the main cytosolic proteases responsible for generating peptides for antigen processing and presentation in the MHC (major histocompatibility complex) class-I pathway. Purified 20S and 26S proteasomes have been widely used to study both specificity and efficiency of antigen processing. Here, we describe the purification of active human 20S and 26S proteasomes from human erythrocytes by DEAE-ion exchange chromatography, ammonium sulfate precipitation, glycerol density gradient centrifugation, and Superose-6 size exclusion chromatography and their characterization using fluorogenic substrates and specific inhibitors.
Molecular tools to assess the diversity and density of denitrifying bacteria in their habitats
2007
Publisher Summary This chapter describes the molecular tools to assess the diversity and density of denitrifying bacteria in their habitats. Genome sequencing and metagenomic projects might even provide new denitrification gene sequences, which could aid in designing more broad range primers. Most information is obtained by cloning and sequencing the polymerase chain reaction (PCR) amplicons, but a more rapid analysis is achieved using fingerprinting techniques. As all PCR-based analyses, the fingerprinting techniques are subjected to well-known biases introduced by, e.g., DNA extraction procedures, primer selection, and PCR conditions. For denitrifiers, the PCR-RFLP (restriction fragment l…
Increasing voltage gradient electrophoresis of DNA
2007
We developed a method which allows electrophoretic fractionation of DNA in an agarose matrix according to an increasing current gradient, using a previously designed [R. Barbieri, V. Izzo, M.A. Costa, G. Giudice, G. Duro, Anal. Biochem. 212 (1993) 168; M.R. Asaro, V. Izzo, R. Barbieri, J. Chromatogr. A 855 (1999) 723] voltage gradient apparatus. This method allows the separation of different DNA fragments by increasing the distances of the components fractionated in the gel, revealing small differences in the length of different DNA components.
A method for eluting DNA in a wide range of molecular weights from agarose gels
1991
We have developed a simple and rapid method for recovering DNAs of a wide range of molecular weights from agarose gels. A DNA-containing gel slice is placed on a Parafilm sheet in the center of a circular (positive) electrode and covered with a drop of buffer, while a linear (negative) electrode is placed on the top of the gel and driven about 1 mm into the gel itself. When a continuous current is applied, the DNA migrates into the buffer toward the circular electrode. We have obtained almost total recovery of DNAs up to 10 kb in size. Our method may also be used, under appropriate conditions, for higher molecular weight DNAs. The yield and all the biological assays performed on the DNAs ob…
Molecular monitoring of inactivation efficiencies of bacteria during pulsed electric field treatment of clinical wastewater
2008
Aims: The applicability of an alternative wastewater disinfection concept based on the pulsed electric field (PEF) treatment is tested with molecular biology techniques using clinical wastewaters. Methods and Results: Hospital wastewater was treated with the PEF technology. The inactivation efficiencies of bacteria were successfully monitored with real-time polymerase chain reaction (PCR). As the differentiation between living and dead bacterial cells is important for the determination of the disinfection efficiency, propidium monoazide (PMA) was applied. PMA selectively penetrates cells with compromised membranes and intercalates into the DNA inhibiting a subsequent PCR amplification. Th…
Haptoglobintypenbestimmung mittels vertikaler S�ulenelektrophorese in Polyacrylamidgel
1972
Ein kontinuierliches und zwei diskontinuierliche Polyacrylamidgelelektrophoreseverfahren zur Haptoglobintypendifferenzierung werden beschrieben und die Vorzuge dieser Technik den mit der Starkegelelektrophorese zu gewinnenden Ergebnissen gegenubergestellt. Alle 3 Verfahren sind fur die Routinediagnostik geeignet. Die kontinuierliche PAA-Gelelektrophorese bedeutet zwar eine geringe Vereinfachung der Methodik, sie ist aber den Disk-Verfahren hinsichtlich der Trennungsergebnisse des Hp-Bandenspektrums unterlegen.