RESUMEN
BACKGROUND: Filamentous bacteriophage are used as general cloning vectors as well as phage display vectors in order to study ligand-receptor interactions. Exposure to biphasic chloroform-water interface leads to specific contraction of phage, to non-infective I- or S-forms. RESULTS: Upon exposure, phage were inactivated (non-infective) at methanol, ethanol and 1-propanol concentrations inversely dependent upon alcohol hydrophobicity. Infectivity loss of phage at certain concentrations of 1-propanol or ethanol coincided with changes in the spectral properties of the f1 virion in ultraviolet fluorescence and circular dichroism studies. CONCLUSIONS: The alcohols inactivate filamentous phage by a general mechanism--solvation of coat protein--thereby disrupting the capsid in a manner quite different from the previously reported I- and S-forms. The infectivity retention of phagemid pG8H6 in 99% acetonitrile and the relatively high general solvent resistance of the phage strains studied here open up the possibility of employing phage display in non-aqueous media.
Asunto(s)
Bacteriófagos/química , Bacteriófagos/crecimiento & desarrollo , Técnicas Químicas Combinatorias/métodos , Solventes/farmacología , 1-Propanol/farmacología , Acetonitrilos/farmacología , Bacteriófagos/efectos de los fármacos , Dicroismo Circular , Medios de Cultivo , Etanol/farmacología , Concentración de Iones de Hidrógeno , Metanol/farmacología , Espectrometría de Fluorescencia , TemperaturaRESUMEN
Swedish soil isolates biochemically classified as Bacillus thuringiensis subsp. israelensis were further examined for genetic diversity by multilocus enzyme electrophoresis (MLEE), random amplified polymorphic DNA analysis (RAPD), pulse field gel electrophoresis (PFGE), and Southern blotting, and were compared with reference strains. All the tested strains belonging to the Bt. israelensis serotype H14 were found to be identical, as judged from the RAPD analysis. MLEE analysis gave a similar result; only one H14 strain was found to differ from the remaining H14 strains by one null allele. PFGE analysis confirmed a very close relationship between the H14 strains but revealed an SfiI restriction fragment of variable size. Southern blot analyses were carried out with probes for the chromosomally encoded flagellin gene(s) and the plasmid-encoded mosquitocidal toxins. All probes gave similar hybridization patterns in the H14 strains. The mosquito toxin probes hybridized only to the H14 strains, except for one probe hybridizing to strain 6:3, which was originally isolated from the same soil sample as strains 6:11 and 6:12. Because the RAPD, MLEE, and PFGE analyses showed that strain 6:3 appears to be unrelated to strains 6:11 and 6:12, the presence of a mosquito toxin sequence in strain 6:3 may suggest that gene transfer has occurred.
Asunto(s)
Bacillus thuringiensis/genética , Toxinas Bacterianas , Variación Genética , Genoma Bacteriano , Microbiología del Suelo , Animales , Bacillus thuringiensis/clasificación , Bacillus thuringiensis/fisiología , Toxinas de Bacillus thuringiensis , Proteínas Bacterianas/genética , Southern Blotting , Electroforesis/métodos , Electroforesis en Gel de Campo Pulsado , Endotoxinas/genética , Enzimas/análisis , Flagelina/genética , Proteínas Hemolisinas , Control Biológico de Vectores , Técnica del ADN Polimorfo Amplificado AleatorioRESUMEN
The stability of the filamentous phages derived from phagemid pG8H6 has been examined in a range of solvents and solvent mixtures. The results show an enhanced capacity to infect E. coli after exposure to various organic solvent-water mixtures. The dependence of stability upon solvent hydrophobicity was demonstrated. Furthermore, conditions have been identified which should allow the application of phage display libraries based upon pG8H6 in organic media.
