Process for determination of a complete or a partial contents of very short sequences in the samples of nucleic acids connected to the discrete particles of microscopic size by hybridization with oligonucleotide probes

摘要

Determination of the formula of genomic DNA, i.e. genome sequencing, by a hybridization with oligonucleotide probes (YU Patent Application 570/87) envisages the use of 100000 oligonucleotide probes and the same number of hybridizati­ons with 6000000 of addressed sample-clones on filters in order to determine contents of oligonucleotide sequences in each clone. The process presents improvements in preparati­on of samples for hybridization and improvements which enable one to follow gene expression by determining partial or complete fragment sequences of genomic DNA, mRNA or cDNA. By binding fragments of genomic DNA to discrete particles (DP) of a microscopic size which are recognizable in a step of reading experimental image, the necessity for addressed samples on filters is dispensed with and this drastically reduces automatical-robotical component of the process and allows miniaturization of the entire method from a level of industrial installation to the level of laboratory instrument. Processes for binding DNA fragments to DPs recognizable in common reactions, allow elimination of cloning, i.e. DNA amplification in the host cells and, in a process of library forming, need for formation of 6000000 addressed samples in any one of the phases of a process for sequencing by hybridization.