[Detection of single base polymorphism in p53 gene by ligase detection reaction and rolling circle amplification on microarrays].

We combined three modern technologies of single base polymorphism detection in human genome: ligase detection reaction, rolling circle amplification and IMAGE hydro-gel microarrays. Polymorphism in target DNA was tested by selective ligation on microarray. Product of the ligase reaction was determined in microarray gel pads by rolling circle amplification.

Identification of rifampin-resistant Mycobacterium tuberculosis strains by hybridization, PCR, and ligase detection reaction on oligonucleotide microchips.

Three new molecular approaches were developed to identify drug-resistant strains of Mycobacterium tuberculosis using biochips with oligonucleotides immobilized in polyacrylamide gel pads. These approaches are significantly faster than traditional bacteriological methods. All three approaches-hybridization, PCR, and ligase detection reaction--were designed to analyze an 81-bp fragment of the gene rpoB encoding the beta-subunit of RNA polymerase, where most known mutations of rifampin resistance are located.

Detection of rifampicin-resistant Mycobacterium tuberculosis strains by hybridization and polymerase chain reaction on a specialized TB-microchip.

Two alternative methods for identification of rifampicin-resistant strains of Mycobacterium tuberculosis on biological microchips are developed. The methods are based on detection of point mutations and other rearrangements in the rpoB gene region determining rifampicin resistance. Hybridization on TB-microchip detects 30 mutant variants of DNA in rifampicin-resistant strains (about 95% of all resistant forms).

Integration of multiple PCR amplifications and DNA mutation analyses by using oligonucleotide microchip.

We have developed a method for parallel independent on-chip amplification and the following sequence variation analysis of multiple DNA regions directly using microchip with an array of nanoliter gel pads containing specific sets of tethered primers. The method has three key features. First, DNA to be amplified is enriched at gel pads by its hybridization with immobilized primers.

PCR amplification on a microarray of gel-immobilized oligonucleotides: detection of bacterial toxin- and drug-resistant genes and their mutations.

PCR amplification on a microarray of gel-immobilized primers (microchip) has been developed. One of a pair of PCR primers was immobilized inside a separate microchip polyacrylamide porous gel pad of 0.1 x 0.

GABA-induced changes of the tissue-specific peptide pool of white rat brain.

Internasal administration of gamma-aminobutyric acid (GABA) induced prolonged behaviour changes and the appearance of three new compounds absent in the brain extracts of control rats. Two peptides associated with GABA administration were isolated and sequenced: Thr-Tyr-Thr-Phe, which corresponds to a gamma-immunoglobulin segment, and Val-Leu, which is present in a great number of proteins, hence its precursor could not be established. The third compound was not amenable to the Edman degradation technique.

Peptides from bovine brain: structure and biological role.

Fractionation of bovine brain extracts followed by automatic Edman sequencing of individual components resulted in identification of 107 endogenous peptides formed from functional proteins (haemoglobin, myelin basic protein, cytochrome c oxidase, etc) or unknown precursors. Several of the newly identified brain peptides demonstrate different types of biological activity; some of the substances show considerable overlap with the known biologically active peptides. It is suggested that these peptides should participate in regulation of extracellular and intracellular biochemical processes.

Cytotoxic activity of acetylcholine receptor ligands.

Acetylcholine receptor ligands were studied for cytotoxicity in K562 human erythroid leukemia tumor cells. Cytotoxicity of carbachol, an agonist of acetylcholine receptors, atropine, an antagonist of muscarinic acetylcholine receptor, neurotoxin 11 (NT II) from Naja naja oxiana cobra venom and tubocurarine, antagonists of acetylcholine receptor of nicotinic type was exhibited in the 10-7-10-5 M concentration range. Several cytolytic processes, two for carbachol and three for other ligands, corresponding to different concentrations of each ligand were detected.

Both neurotoxin II from venom of Naja naja oxiana and its endogenous analogue induce apoptosis in tumor cells.

Both neurotoxin II (NT II from venom of Naja naja oxiana) and 20-30 kDa proteins partially purified from pig brain (NTlm) cross-reacting with antibodies to NT II were cytotoxic for L929 and K562 tumor cells at concentrations of 10(-6)-10(-8) M. Modification of Cys residues significantly reduced cytotoxicity of NT II. Both NT2 and NTlm induced apoptosis in L929 and K562 cells.