National Institute of Advanced Industrial Science and Technology

Kankyo Engineering Co., Ltd.

Nippon Steel & Sumikin Eco-tech Corporation

Hidenori Tani

Kazunori Nakamura

Takahiro Kanagawa

Shinya Kurata

Satoshi Tsuneda

Naohiro Noda

Masaki Torimura

Hiroaki Sato

The present invention provides means for evaluating harmfulness of a chemical substance before occurrence of cell death, that is, more quickly and sensitively compared to conventional methods wherein the remaining viable cell count is determined using a reductive coloring reagent after a period of time required for occurrence of cell death due to the chemical substance. A double-stranded RNA probe comprising an RNA strand labeled with a fluorescent dye A that emits fluorescence, and an RNA strand labeled with a fluorescent dye B that quenches emission from a fluorescent dye in the vicinity thereof, wherein the fluorescence is quenched in a double-stranded state due to occurrence of fluorescence resonance energy transfer (FRET) between the two kinds of fluorescent dyes. When the probe is introduced into a cell and the cell is in a normal state, the double-stranded RNA is quickly degraded by activity of intracellular ribonuclease to cause cancellation of the FRET state, allowing fluorescence emission of the fluorescent dye A. On the other hand, in cases where the cell is harmfully influenced, activity of intracellular ribonuclease is suppressed, so that the double-stranded RNA remains undegraded, and the fluorescent dye A remains quenched due to the influence of the fluorescent dye B. By detecting a decrease in the degradation rate of intracellular RNA based on this principle, harmfulness of a chemical substance can be quickly and sensitively evaluated before occurrence of cell death.

Method for measuring in vivo inhibition of intracellular RNase

[Problems] To provide a novel mixture for assaying a target nucleic acid, characterized by enabling a nucleic acid assay while: 1) requiring no step of diluting the target nucleic acid; 2) requiring no procedure of changing a probe concentration depending on a concentration of the target nucleic acid. [Means for Solving Problems] 1) A mixture which comprises one internal standard nucleic acid and two nucleic acid probes labeled with a fluorescent dye; 2) a mixture for measuring Km value which comprises one internal standard nucleic acid having a partial mutation and one nucleic acid probe labeled with a fluorescent dye; 3) a mixture which comprises one internal standard nucleic acid and one double nucleic acid probe labeled with two fluorescent dyes; and a method for assaying a nucleic acid by making use thereof.

Mixtures for assaying nucleic acid, novel method of assaying nucleic acid with the use of the same and nucleic acid probe to be used therefor

To provide a novel mixture for assaying a target nucleic acid, characterized by enabling a nucleic acid assay while: 1) requiring no step of diluting the target nucleic acid; 2) requiring no procedure of changing a probe concentration depending on a concentration of the target nucleic acid. 1) A mixture which comprises one internal standard nucleic acid and two nucleic acid probes labeled with a fluorescent dye; 2) a mixture for measuring Km value which comprises one internal standard nucleic acid having a partial mutation and one nucleic acid probe labeled with a fluorescent dye; 3) a mixture which comprises one internal standard nucleic acid and one double nucleic acid probe labeled with two fluorescent dyes; and a method for assaying a nucleic acid by making use thereof.

Growing community of inventors

Tokyo, Japan

Hidenori Tani