Solvolysis of 2'-deoxyribonucleosides and oligo-2'-deoxyribonucleotides in aqueous pyrrolidine or ammonia solutions.

To assess the feasibility of high-temperature aminolysis of deoxyribooligonucleotides containing rare bases as a method to determine their base sequence, the 2'-βD-deoxyribosides of 5-bromouracil, 2-aminopurine, uracil, adenine, cytosine, 5-methylcytosine, hypoxanthine, N-methyladenine, N-ethylcytosine, and guanine were compared as to their rate of degradation in 0.5 M aqueous pyrrolidine at 110 °C, conditions used earlier in the analysis of oligonucleotides containing only the canonical bases. The reaction mixtures were analyzed by chromatography on Zorbax XDB-CN and UV absorption spectroscopy.

Residue interactions affecting the deprotonation of internal guanine moieties in oligodeoxyribonucleotides, calculated by FMO methods.

The effect of vicinal molecular groups on the intrinsic acidity of a central guanine residue in short single-stranded DNA models and the potentials exerted by the backbone and the nucleobases on the leaving proton were determined by the fragment molecular orbital (FMO) method, in terms of quantum descriptors (QDs) and pair interaction interfragment decomposition analysis (PIEDA). The acidity of the central guanine moiety decreased with increasing oligonucleotide length, in response to changes by less than 1 eV in the ionization potential, global softness, electrophilicity index, and electronegativity descriptors. The differences in these descriptors were majorly interpreted in terms of the electrostatic influence of the negative charges residing on the backbone of the molecule.

Nanosilver gel as an endodontic alternative against Enterococcus faecalis in an in vitro root canal system in Mexican dental specimens.

Nanotechnology has become a research area with promising results for technological innovation. Endodontics can benefit from this field of research by increasing the success rate of the treatment, which currently ranges between 86% and 98% and has varied very little over the years. One of the causes of endodontic treatment failure is based on the presence of Enterococcus faecalis.

czcD gene from Bacillus megaterium and Microbacterium liquefaciens as a potential nickel-vanadium soil pollution biomarker.

Metals are among the most prevalent pollutants released into the environment. For these reasons, the use of biomarkers for environmental monitoring of individuals and populations exposed to metal pollution has gained considerable attention, offering fast and sensitive detection of chemical stress in organisms. There are different metal resistance genes in bacteria that can be used as biomarkers, including cation diffusion facilitators carrying metal ions; the prototype is the cobalt-zinc-cadmium transporter (czcD).

High-temperature solvolysis of 2'-deoxyribonucleosides in aqueous amine solutions.

Degradation of 2'-deoxyribonucleosides in 0.5 M aqueous pyrrolidine at 110 °C proceeds at different rates, ordered as deoxyuridine > deoxyadenosine > deoxycytidine > deoxyguanosine ≫ deoxythymidine. Deoxyadenosine degradation produces the free base, adenine, while deoxycytidine by deamination produces deoxyuridine, and then uracil.

Protonation of Nucleobases in Single- and Double-Stranded DNA.

Single-stranded model oligodeoxyribonucleotides, each containing a single protonatable base-cytosine, adenine, guanine, or 5-methylcytosine-centrally located in a background of non-protonatable thymine residues, were acid-titrated in aqueous solution, with UV monitoring. The basicity of the central base was shown to depend on the type of the central base and its nearest neighbours and to rise with increasing oligonucleotide length and decreasing ionic strength of the solution. More complex model oligonucleotides, each containing a centrally located 5-methylcytosine base, were comparatively evaluated in single-stranded and double-stranded form, by UV spectroscopy and high-field NMR.

Use of alternative alkali chlorides in RT and PCR of polynucleotides containing G quadruplex structures.

Several alkali chlorides were compared for their use in reverse transcription (RT) and PCR of different types of nucleic acid templates. On a test region of biological DNA incapable of forming G quadruplex (G4) structures, Taq DNA polymerase showed similar PCR performance with 50 mM KCl, CsCl, LiCl, and NaCl. In contrast, on a synthetic model polydeoxyribonucleotide prone to G4 formation, good PCR amplification was obtained with 50 mM CsCl, but little or none with LiCl or KCl.

Expression Changes in Metal-Resistance Genes in Microbacterium liquefaciens Under Nickel and Vanadium Exposure.

Microbacterium liquefaciens MNSH2-PHGII-2 is a nickel-vanadium-resistant bacterium isolated from mine tailings located in Guanajuato, Mexico. In PHGII liquid media, M. liquefaciens has the ability to remove 29.

PCR amplification of GC-rich DNA regions using the nucleotide analog N4-methyl-2'-deoxycytidine 5'-triphosphate.

GC-rich DNA regions were PCR-amplified with Taq DNA polymerase using either the canonical set of deoxynucleoside triphosphates or mixtures in which the dCTP had been partially or completely replaced by its N4-methylated analog, N4-methyl-2'-deoxycytidine 5'-triphosphate (N4me-dCTP). In the case of a particularly GC-rich region (78.9% GC), the PCR mixtures containing N4me-dCTP produced the expected amplicon in high yield, while mixtures containing the canonical set of nucleotides produced numerous alternative amplicons.

Identification of Bacillus megaterium and Microbacterium liquefaciens genes involved in metal resistance and metal removal.

Bacillus megaterium MNSH1-9K-1 and Microbacterium liquefaciens MNSH2-PHGII-2, 2 nickel- and vanadium-resistant bacteria from mine tailings located in Guanajuato, Mexico, are shown to have the ability to remove 33.1% and 17.8% of Ni, respectively, and 50.

Determination of pKa values for deprotonable nucleobases in short model oligonucleotides.

The deprotonation of ionizable nucleobases centrally placed in short model oligonucleotides was examined under different physical conditions, using UV absorption spectroscopy. The oligonucleotide sequences were designed so that only the central base would be ionized over the pH range examined. pKa values of 9.

Hybridisation of N4-methylcytosine-containing amplicons on DNA microarrays.

5'-Cy5-labelled PCR amplicons containing the analogue base, N(4)-methylcytosine, instead of cytosines were compared in microarray hybridisation experiments with the corresponding amplicons containing the canonical set of bases, with respect to the intensity of the fluorescence signal obtained, and cross hybridisation to non-corresponding probes. In general, higher hybridisation temperatures resulted in reduced signal intensities, particularly in the case of the N(4)-methylcytosine containing amplicons. At the lower hybridisation temperatures tested (40 °C, 30 °C), these modified amplicons gave about equal or stronger fluorescence signal than the corresponding regular amplicons.

Capacity of N4-methyl-2'-deoxycytidine 5'-triphosphate to sustain the polymerase chain reaction using various thermostable DNA polymerases.

The dCTP analog N4-methyl-2'-deoxycytidine 5'-triphosphate (N4medCTP) was evaluated for its performance in the polymerase chain reaction (PCR). Using the HotStart Taq DNA polymerase with a standard thermal protocol, test segments 85 or 200 bp long were amplified equally well using dCTP or N4medCTP:dCTP mixtures ranging in molar ratio from 3:1 to 10:1, while complete replacement of dCTP by N4medCTP gave clearly lower amplicon yields and higher Cq values. Comparable yields with N4medCTP or dCTP were achieved only by using a slowdown protocol.

Comparison of different amines for the one-lane sequence determination of 5'-end-labeled oligodeoxyribonucleotides by chemical cleavage.

Hot aqueous solutions of a wide variety of aliphatic amines bring about efficient cleavage of 5'-(32)P-labeled oligodeoxyribonucleotides. Electrophoretic resolution of the product mixtures produces a radioactivity pattern that can be analyzed in terms of relative band intensities and band separations to deduce the base sequence of the original oligonucleotide. Amines differ in their overall reactivity with respect to the oligonucleotide as well as in their preference for reaction with the various heterocyclic bases.