Adsorption of extracellular chromosomal DNA and its effects on natural transformation of Azotobacter vinelandii.
نویسندگان
چکیده
To better understand the influence of environmental conditions on the adsorption of extracellular chromosomal DNA and its availability for natural transformation, the amount and conformation of adsorbed DNA were monitored under different conditions in parallel with transformation assays using the soil bacterium Azotobacter vinelandii. DNA adsorption was monitored using the technique of quartz crystal microbalance with dissipation (QCM-D). Both silica and natural organic matter (NOM) surfaces were evaluated in solutions containing either 100 mM NaCl or 1 mM CaCl(2). The QCM-D data suggest that DNA adsorbed to silica surfaces has a more compact and rigid conformation in Ca(2+) solution than in Na(+) solution and that the reverse is true when DNA is adsorbed to NOM surfaces. While the amounts of DNA adsorbed on a silica surface were similar for Ca(2+) and Na(+) solutions, the amount of DNA adsorbed on an NOM-coated surface was higher in Ca(2+) solution than in Na(+) solution. Transformation frequencies for dissolved DNA and DNA adsorbed to silica and to NOM were 6 x 10(-5), 5 x 10(-5), and 2.5 x 10(-4), respectively. For NOM-coated surfaces, transformation frequencies from individual experiments were 2- to 50-fold higher in the presence of Ca(2+) than in the presence of Na(+). The results suggest that groundwater hardness (i.e., Ca(2+) concentration) will affect the amount of extracellular DNA adsorbed to the soil surface but that neither adsorption nor changes in the conformation of the adsorbed DNA will have a strong effect on the frequency of natural transformation of A. vinelandii.
منابع مشابه
Interactions between dissolved natural organic matter and adsorbed DNA and their effect on natural transformation of Azotobacter vinelandii.
To better understand gene transfer in the soil environment, the interactions between dissolved natural organic matter (NOM) and chromosomal or plasmid DNA adsorbed to silica surfaces were investigated. The rates of NOM adsorption onto silica surfaces coated with DNA were measured by quartz crystal microbalance (QCM) and showed a positive correlation with carboxylate group density for both soil ...
متن کاملTransformation of Azotobacter vinelandii with plasmid DNA.
Azotobacter vinelandii cells can be transformed at high frequencies with the broad-host-range plasmids pRK2501, RSF1010, and pGSS15, using a modification of the procedure developed by Page and von Tigerstrom (J. Bacteriol. 139:1058-1061, 1979) for chromosomal DNA-mediated transformation. The frequency of transformation per microgram of plasmid DNA per viable cell with pRK2501 and pGSS15 was abo...
متن کاملGene dosage analysis in Azotobacter vinelandii.
For more than a decade, Azotobacter vinelandii has been considered a polyploid bacterium on the basis of physical studies of chromosome size and DNA content per cell. However, as described in the present work, many genetic operations can be performed in A. vinelandii without the constraints expected in a polyploid bacterium: (i) reversion of transposon-induced mutations is usually associated wi...
متن کاملHeat sensitivity of Azotobacter vinelandii genetic transformation.
Heating competent Azotobacter vinelandii at 37 or 42 degrees C resulted in a total loss of competence with no loss of viability. The transformation process was relatively insensitive to heating at either temperature once DNase-resistant DNA binding was nearly complete. Although competent and 42 degrees C-treated cells bound equivalent amounts of [32P]DNA in a DNase-resistant state, no donor DNA...
متن کاملOptimal conditions for transformation of Azotobacter vinelandii.
Optimal transformation of Azotobacter vinelandii OP required a 20-min incubation of the competent cells with deoxyribonucleic acid at 30 degrees C in buffer (pH 6.0 to 8.0) containing 8 mM magnesium sulfate. Nitrogen-fixing transformants of nitrogen fixation-deficient recipients could be plated immediately on selective medium, but transformants acquiring rifampin and streptomycin resistance req...
متن کاملذخیره در منابع من
با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید
برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید
ثبت ناماگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید
ورودعنوان ژورنال:
- Applied and environmental microbiology
دوره 76 13 شماره
صفحات -
تاریخ انتشار 2010