[The efficacy of hepatotropic agent Remaxol in oncological patients with postoperative liver dysfunction].

The study of the efficiency and safety of the hepatotropic drug Remaxol in oncological patients with minimal, subclinical manifestations of liver dysfunction in the postoperative period was performed. It is shown that using of Remaxolcontributes to leveling the biochemical liver dysfunction: decrease of the concentration of total bilirubin, a decrease serum activity of lactate dehydrogenase, alanine aminotransferase, aspartate aminotransferase, and alkaline phosphatase. The unwanted effects of Remaxol were not identified.

[Bioluminescent reporters to identify gene allelic variants].

The method of single nucleotide polymorphism identification based on primer extension reaction (PEXT) with the following bioluminescent solid-phase microassay was developed. The recombinant Ca2+-regulated photoprotein obelin and coelenterazine-dependent luciferase Renilla muelleri were used as reporters. Factor V Leiden polymorphism 1691 G-->A (R506Q) of human F5 gene genotyping was used for investigation.

[Cloning of genes, purification and properties investigation of recombinant DNA ligases from the thermophilic archaeon Pyrococcus abyssi and Methanobacterium thermoautotrophicum].

The genes encoding of DNA ligases from the thermophilic archaeon Pyrococcus abyssi (PabDNA ligase) and Methanobacterium thermoautotrophicum (MthDNA ligase) were cloned and expressed in Escherichia coli. The activity of purified enzymes was studied by ligation of two oligonucleotides, one of which had preformed hairpin structure. In the used system the maximal output of reaction products for both DNA ligases was observed near 70 degrees C that is explained by substrate thermostability.

[Effect of sweet pepper, enriched with selenium, on the transplantable Ehrlich carcinoma in mice].

Effect of sweet pepper enriched and non-enriched with Se on Ehrlich carcinoma growth at Balb/c mice was investigated. 1000 mg/kg of both preparations of sweet pepper (per os administration for mice of 9 months age) inhibited an early stage of tumour growth on 42-62% and 37-65% respectively. No effect on tumour growth was registered for 4 months mice.

[Oligonucleotide derivatives in the nucleic acid hybridization analysis. II. Isothermal signal amplification in process of DNA analysis by minisequencing].

The isothermal amplification of reporter signal via limited probe extension (minisequencing) upon hybridization of nucleic acids has been studied. The intensity of reporter signal has been shown to increase due to enzymatic labeling of multiple probes upon consecutive hybridization with one DNA template both in homophase and heterophase assays using various kinds of detection signal: radioisotope label, fluorescent label, and enzyme-linked assay. The kinetic scheme of the process has been proposed and kinetic parameters for each step have been determined.

[Oligonucleotide derivatives in the nucleic acid hybridization analysis. I. Covalent immobilization of oligonucleotide probes onto the nylon].

The features of UV-induced immobilization of oligonucleotides on a nylon membranes and the effectiveness of enzymatic labeling of immobilized probes at heterophase detection of nucleic acids are studied. Short terminal oligothymidilate (up to 10 nt) sequences are suggested to attach to the probe via a flexible ethylene glycol based linker. The presence of such fragment enhances the intensity of immobilization and reduces UV-dependent degradation of the targeted (sequence-specific) part of the probe by reducing the dose needed for the immobilization of DNA.

[Bent dsDNA with defined geometric characteristics in terms of complexes of bridged oligonucleotides].

An opportunity of designing nontypical double-stranded DNA structures containing nonnatural inserts in a regular nucleotide DNA sequence has been investigated. The looped nucleotide inserts on the basis of adenylates and thymidilates, and nonnucleotide inserts on the basis of phosphodiesters of diethyleneglycol, 1,10-decanediol, and 3-hydroxy-2-hudroxymethyltetrahydrofuran were introduced into the backbone of a 32-mer native DNA duplex. These inserts formed the internal loops in the modified double-stranded DNA fragments which were shown to lead to bending of the linear duplex structure by 16 to 83 degrees.

[A highly sensitive and rapid method for the detection of DNA fragments using the photoprotein obelin as a reporter].

The recombinant Ca2+-activated photoprotein obelin was used as a reporter protein in a solid-phase bioluminescent hybridization DNA assay. Oligonucleotide probes were immobilized on the surface of a fine-grained polymer or microbiological plates of different types. A 30-mer oligonucleotide or its derivative with a biotin residue on the 3'-terminus, as well as a denatured double-stranded PCR fragment of the hepatitis C virus with the sequence of the 30-mer oligonucleotide, was used as a DNA template.

[Enhancement of hybridization analysis efficacy by means of controlled fragmentation of analyzed DNA].

The influence of fragmentation of analyzed DNA amplicon on the efficacy of specific sequence detection by means of heterophase hybridization analysis was investigated. The detection of DNA sequence was carried out colorimetrically after introduction of biotin label into the oligonucleotide probe immobilized on a solid support upon its limited elongation in the complex with the analyzed DNA using Taq polymerase. Two simple and reproducible approaches to DNA analyte fragmentation were suggested.

[Determination of Herpes simplex virus DNA with the use of solid-phase methods for the detection of PCR products].

To determine DNA of herpes simplex virus (HSV), types 1 and 2, the polymerase chain reaction (PCR) method was developed with the subsequent detection of amplification products by means of electrophoresis or the molecular hybridization of nucleic acids (MHNA). Two variants of MHNA have been compared: hybridization in the solution of a biotinylated probe with digoxigenin-labeled PCR with the subsequent sorption of hybridization complexes onto streptavidin-covered plates and solid-phase hybridization of digoxigenin-labeled PCR with a biotinylated probe. Effective hybridization was observed after the denaturation of targets at 95 degrees C in the solution of 50 mM NaOH, but not in neutral solutions.

[Cleavage of RNA in hybrid duplexes by E. coli ribonuclease H. II. Substrate properties of nucleotides containing non-nucleotide linkers].

The 20-mer bridged oligodeoxynucleotides containing short oligomers joined by the hexamethylenediol and hexaethylene glycol linkers were shown to form complementary DNA/DNA and RNA/DNA complexes whose thermostability depends on the length and number of the nonnucleotide linkers. Hybrid complexes of the bridged oligonucleotides proved to be substrates for the E. coli ribonuclease H.

[Interaction of short nucleotide derivatives with nucleic acids. IV. Modification of DNA by an alkylating tetranucleotide reagents in the presence of effectors in perfect and imperfect complexes].

It was demonstrated that any mismatches in a complex formed by an ssDNA target and a tetranucleotide at 25 or 37 degrees C can be discriminated by alkylating the DNA with a tetranucleotide carrying a 4-[N-methyl-N-(2-chloroethyl)]aminobenzylethylamine residue at the 5'-terminal phosphate in the presence of a pair of flanking effectors, octanucleotide di-N-(2-hydroxyethyl)-phenazinium derivatives. The discrimination factor (ratio of the extent of the target modification in the perfect and mismatch-containing complexes) for a single mismatch in the tetranucleotide binding site at 25 degrees C varied between 4 and 500 depending on the type of mismatch and its location in the complex and exceeded 400 at 37 degrees C for all the investigated mismatches. The DNA target modification by the alkylating derivative of the 3'-estrone ester of tetranucleotide pCAGX (mean = C, T, A or G) was selective in the presence of a pair of hydrophobic effectors, octanucleotide 5'-cholesteryl-3'-phenazinium derivatives.

[Nucleic acids interactions with short oligonucleotide derivatives. II. Tandem of short oligonucleotides as highly sensitive system for identification of single base substitutions in target DNA].

A new approach for modification of target DNAs with tandems of derivatives of short oligonucleotides was suggested that allows highly selective modification of perfect duplexes only. At physiological temperatures, the efficiency of DNA modification by a dodecanucleotide alkylating agent was demonstrated to be the same for both perfect and mismatch-containing duplexes, whereas the tetranucleotide reagent in the presence of two flanking effectors alkylated with high selectivity the target DNA in the perfect duplex only.

[Interaction of derivatives of short oligonucleotides with nucleic acids. I. Effect of various types of effectors on alkylation of DNA-targets].

It was shown that the tandem of the derivatives of short oligonucleotides efficiently and site specifically interacts with target 20 base deoxyribonucleotide (M). It was demonstrated that the very low hybridization ability of tetranucleotide (D) and its 3'-cholesterol and 3'-estrone esters (D-ChS and D-EsS, respectively) increases significantly in the presence of the effectors: octanucleotides (E1 and E2), and their 5',3'-diphenazinium (Phn-E1-Phn and Phn-E2-Phn) and 5'-cholesteryl-3'-phenazinium (ChS-E1-Phn and ChS-E2-Phn) derivatives, which flank them on the target strand. The influence of the effectors on the interaction of the target M with tetranucleotide D or its alkylating derivatives (RCl-D) increases in a series E1 + E2 < ChS-E1-Phn + ChS-E2-Phn < Phn-E1-Phn + Phn-E2-Phn.