RÉSUMÉ
The site-specific endonuclease AbaI was isolated and purified to functional purity from the soil nitrogen-fixing bacterium Azospirillum brasilense UQ 1796. Purification included successive chromatography on columns with phosphocellulose, heparin-Sepharose, and hydroxyapatite. The purified enzyme recognizes the palindromic DNA sequence 5'-T decreases ATCA-3' and cleaves it as shown by the arrow. The isolated enzyme belongs to class II restriction endonuclease and is an isoschizomer of endonuclease BclI. The enzyme of AbaI is active at 26-56 degrees C. The optimal temperature is 48 degrees C and the optimal buffer is LRB.
Sujet(s)
Azospirillum brasilense/enzymologie , DNA restriction enzymes/isolement et purification , DNA restriction enzymes/métabolisme , Type II site-specific deoxyribonuclease/métabolisme , Cellulose/analogues et dérivés , Chromatographie , ADN/métabolisme , ADN viral/métabolisme , Durapatite , Masse moléculaire , Agarose/analogues et dérivés , Spécificité du substrat , TempératureRÉSUMÉ
A critical step in any epidemiologic research concerning nosocomial infections is the precise identification of the responsible pathogen. The present work utilized a molecular approach-plasmids identification, restriction lengght polymorphism DNA analysis and random amplified polymorphic DNA for the characterization of 6 nosocomial outbreaks due to 52 strains of methicillin-resistant staphylococcus aureus (MRSA). In these episodes, the clinic-epidemiologic and phenotipic analysis (antibiotype) pointed to a nosocomial infection. Through molecular analysis it was possible to establish in a very precise way, clonality due to MRSA strains in 2 of the studied outbreaks; the same type of analysis allowed to eliminate a MRSA clonal origin in the remainder 4 episodes. The antibiogram was not an useful analytic tool due to its poor discriminatory power. Also, through a PCR procedure, it was possible to identify the presence of the gen mecA in every of the 52 MRSA strains studied
Sujet(s)
Staphylococcus aureus/ultrastructure , Biologie moléculaire , Infection croisée/anatomopathologie , Infections à staphylocoques/épidémiologie , Techniques in vitro , Staphylococcus aureus/effets des médicaments et des substances chimiques , Staphylococcus aureus/pathogénicité , Résistance microbienne aux médicaments/physiologie , Tests de sensibilité microbienne , Résistance à la méticilline/physiologie , DNA restriction enzymes/isolement et purification , Infection croisée/épidémiologie , PlasmidesRÉSUMÉ
El sistema de restrición-metilación (RMSII) Pvull de P. vulgaris fue clonado y expresado con altos niveles en E. coli. Se estudió el efecto de la alta expresión de restrictasa y metilasa en diferentes cepas de E, coli, resultando la cepa HB101 (mrcB) la única eficientemente transformable con plasmidios portadores de RMSII Pvull. Se aplicó un nuevo procedimiento para la purificación de la restrictasa Pvull recombinante utilizando cromatografía de intercambio iónico. Los métodos empleados permitieron la obtención de grandes cantidades de restrictasa Pvull, libre de exonucleasas y endonucleasas inespecíficas
Sujet(s)
Clonage moléculaire , DNA restriction enzymes/isolement et purification , Expression des gènes , Proteus vulgarisRÉSUMÉ
The isolation and characterization of a restriction endonuclease from a thermophilic strain of Bacillus is described. The enzyme recognizes the palindromic sequence 5'...GGCC...3' as determined by PEI-cellulose chromatography of pancreativc DNAse and snake venom phosphodieterase digestion products of labelled DNA fragments, analysis of restriction digests and direct sequence analysis. The enzyme, denominated BspBR, is an isoschizomer of HaeIII and BspRI