Shimadzu Corporation

Naoyuki Nishimura

Yoshinari Shirasaki

Tetsuo Ohashi

Shigeru Fukushima

Jun Tada

Koichi Yamagata

Hiroko Ozaki

Tomoko Nakayama

Shingo Kato

Hiroshi Tonoike

Reiko Yoshida

The present invention provides a method for examining a foreign matter derived from a living body in quality control for production of various products in order to rapidly identify an individual from whom a living body-derived material contaminated as a foreign matter in products or facilities involved in production of the products was derived, while securing the secret of information on nucleic acid sequences unique to individuals. The method for examining a foreign matter derived from a living body includes identifying an individual from whom a living body-derived material contaminated as a foreign matter in products or facilities involved in production of the products was derived, on the basis of information on sequences of nucleic acid contained in the living body-derived material.

Method of examining foreign matter derived from living body

The present invention is a method for synthesis of nucleic acids to amplify an intended nucleic acid in a region in which a GC content is rich, wherein a polyhydric alcohol and/or ammonium sulfate is present in an amplification reaction solution. According to the present invention, it is possible to amplify nucleic acids in a GC rich region efficiently and directly from a sample such as blood containing lots of PCR inhibitory substances without undergoing a process of isolating and purifying the nucleic acid, even though conducting PCR in the GC rich region tends to be difficult using conventional processes even if purified DNA is used.

Method for synthesis of nucleic acids

An object of the present invention is to establish a method for amplifying an RNA in a biologically-derived sample by preparing a reaction solution in which a nucleic acid synthesis is not inhibited even in the presence of various biologically-derived impurities and as well as to enable a convenient, rapid and highly sensitive analysis of the RNA in the sample. In a method of the present invention, a reaction solution having a pH higher than that of an ordinarily employed reaction solution and/or containing a polyamine and/or a sulfated polysaccharide.

Method for amplification of RNA

If polyamine is caused to be present in the reacting system for the synthesis of nucleic acids to amplify target genes from a sample, the inhibition of the synthesis due to impurities in the sample can be reduced and the target genes can be effectively synthesized.

Enhancement of nucleic acid amplification by the addition of a polyamine

This invention is directed to a novel method for PCR amplification wherein PCR is carried out at a higher pH than the pH widely used in the art. Specifically, the buffer solution is adjusted to pH 9.0 to 11.0 at 25.degree. C. Using the present invention, DNA amplification can be successfully carried out following a simple pretreatment. In the present invention whole blood is mixed with a hypotonic solution so that a selective lysis of red blood cells takes place. The residual leukocytes are then collected. The leukocytes are mixed with a polymerization agent, primers and other necessary reagents and PCR is carried out. When the PCR solution is placed at a high temperature for DNA denaturation, the leukocytes are lysed so that the leukocyte DNA is released and can access the primers and the other necessary reactions for PCR in the solution. Cell membranes and proteins are present in the PCR reaction solution due to the lack of a protein extractive step during the pretreatment. Nevertheless DNA amplification occurs under the presently claimed improved PCR method.

Process and reagent for amplifying nucleic acid sequences

The present invention provides a method of nucleic acid synthesis capable of directly amplifying a gene of interest in a salivary sample without purifying DNAs from the sample. According to the present invention, in a method of nucleic acid synthesis by mixing a salivary sample itself and a gene amplification reaction solution, and then subjecting them to an amplification reaction, the salivary sample is heat-treated or treated with a saccharide-degrading enzyme before the reaction. For example, a salivary sample was heat-treated at 0-40.degree. C. for 1 hour, subsequently added directly into a PCR reaction solution, and then subjected to a PCR. The result demonstrates that, as shown in FIG. 3, the amount of the PCR product increased according as the heat-treatment temperature was elevated, and as a consequence, the PCR product sufficient to be detected in the electrophoresis was produced in each of all cases using different amounts of saliva, by heat-treating the sample at 30.degree. C. or above.

Oligonucleotides (SEQ ID NOs 1-8) selectively hybridizable with a specific gene of Vibro parahaemolyticus, oligonucleotides (SEQ ID NOs 9-13) selectively hybridizable with the LT gene of toxigenic Escherchia coil, oligonucleotides (SEQ ID NOs 14-21) selectively hybrizable with the STh or STp gene of toxigenic Escherchia coil, oligonucleotides (SEQ ID NOs 22-47) selectively hybridizable with the entA, B, C, or D gene of Staphylococcus aureus, or oligonucleotides (SEQ ID NOs 48-53) selectively hybridizable with the entE gene of Staplyloccus aureus are prepared and used as primers for gene amplification to thereby selectively detect only respective microorganisms causing food poisoning.

Oligonucleotides for detecting bacteria and detection method using same

Oligonucleotides (SEQ ID NOs 1-8) selectively hybridizable with a specific gene of Vibro parahaemolyticus, oligonucleotides (SEQ ID NOs 9-13) selectively hybridizable with the LT gene of toxigenic Escherichia coil, oligonucleotides (SEQ ID NOs 14-21) selectively hybrizable with the STh or STp gene of toxigenic Escherichia coil, oligonucleotides (SEQ ID NOs 22-47) selectively hybridizable with the entA, B, C, or D gene of Staphylococcus aureus, or oligonucleotides (SEQ ID NOs 48-53) selectively hybridizable with the entE gene of Staplyloccus aureus are prepared and used as primers for gene amplification to thereby selectively detect only respective microorganisms causing food poisoning.

Growing community of inventors

Kyoto, Japan

Naoyuki Nishimura