# US 5,702,883 A · Methods for detection of mutagens using luminescence gene

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- Number: US 5,702,883 A
- Document ID: US5702883A
- Status: Granted
- Kind: A
- Application number: 08/326,949
- Filed: 1994-10-21
- Granted: 1997-12-30
- Earliest claimed: 1993-10-22
- Assignees: Kabushiki Kaisha Toyota Chuo Kenkyusho
- Inventors: Takao Imaeda; Masana Hirai
- Web page: https://patentgrep.com/patent/US5702883A

## CPC

- C07K 14/47: from mammals
- C12Q 1/025: {for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics (antimicrobial activity C12Q1/18)}
- C12Q 1/6897: involving reporter genes operably linked to promoters
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES

## Abstract

A method for detecting or quantitating a mutagenic substance in a sample includes culturing a host microorganism transformed with a recombinant gene comprising an SOS gene and genes expressing luciferase activity and optionally genes expressing an enzyme which catalyzes the production of a substrate for luciferase, positioned downstream of the SOS gene, in a medium to which the sample is added; and measuring a luminescence generated by expression of the gene expressing luciferase activity. The method is sensitive, accurate and non-time consuming; and gene systems used for said method, i.e., a recombinant gene comprising an SOS gene expressed when a DNA is damaged and a gene expressing luciferase activity positioned downstream of the SOS gene, and a host microorganism transformed with said recombinant gene. Preferably the recombinant gene further comprises a gene expressing an enzyme which catalyses the production of a substrate for the luciferase in the down stream of the SOS gene.

## Claims

### Claim 1

A method for identifying a mutagenic substance, comprising the steps of: culturing Escherichia or Salmonella transformed with recombinant DNA comprising (i) an scherichia or Salmonella SOS gene and (ii) a gene encoding luciferase isolated from Photobacterium phosphoreum, Photobacterium leiognethi, Vibrio splendidus, Vibrio cholerae, Photinus pyralis, Vibrio harveyi or Vibrio fischeria;
wherein said gene encoding luciferase is positioned downstream of said SOS gene such that when said SOS gene is expressed, then said gene encoding luciferase is also expressed;
contacting said culture with a substance to be tested wherein said Escherichia or Salmonella in said culture is provided with a luciferase substrate; and
determining whether said substance is mutagenic by measuring the amount of luminescence in said culture.

### Claim 2 (depends on 1)

The method of claim 1, wherein said method comprises collecting and disrupting said cultured Escherichia or salmonella to release luciferase therefrom and providing said luciferase substrate to said released luciferase.

### Claim 3 (depends on 1)

The method of claim 1, wherein, when said luciferase gene is isolated from Photobacterium leiognethi, Vibrio splendidus, Vibrio cholerase, Vibrio harveyi or Vibrio fischeria, said recombinant DNA further comprises luciferase operon genes encoding a fatty acid reductase, positioned downstream from said SOS gene, so that when said SOS gene is expressed, said fatty acid reductase genes are expressed.