Organic compounds -- part of the class 532-570 series – Organic compounds – Carbohydrates or derivatives
Patent
1997-07-28
1999-02-09
Zitomer, Stephanie W.
Organic compounds -- part of the class 532-570 series
Organic compounds
Carbohydrates or derivatives
435 6, 435 912, 536 243, 536 2432, 935 77, 935 78, C07H 2104, C12Q 168
Patent
active
058696423
DESCRIPTION:
BRIEF SUMMARY
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates to the detection of the genus Pectinatus which is known as beer spoilage bacteria. More particularly, the present invention relates to a gene sequence for detecting Pectinatus cerevisiiphilus, Pectinatus frisingensis and Pectinatus sp. DSM20764, respectively, and to a method for detecting the bacteria by using the sequence.
2. Description of the Related Art
Presently, in a method for testing micro-organisms in the field of beer production and, for identifying the species, the bacteria must be separated via growth cultivation and segregation cultivation, which takes at least seven days. Then the, separated bacteria is propagated, and the identification is conducted by many tests such as observation of the form and Gram stain properties, a catalase test, and presence of sugar consumption. These troublesome tests require many hours and the tests are expensive. In addition to these conventional identification methods, there is a method for identifying species by a hybridization test, in which DNA is extracted from an isolated bacteria and the DNA is fixed on a membrane or other substrate, and detected with the DNA of a standard bacteria as a probe. However, this method takes several days, and it is difficult to obtain the necessary detection sensitivity.
Recently, for detecting a bacteria quickly, there is a method for detecting a living bacteria by an ATP luminescence method using reported in J. Am. Soc. Brew. Chem.: 52(1) 19-23, 1994. But this method is unable to identify the species. Further, for a part of lactic acid bacteria, for example, Lactobacillus brevis, Lactobacillus plantarum and Lactobacillus coryniformis, as disclosed in Japanese Patent Laid-open Publication Nos. 6-46811(1994) and 6-311894(1994), it is possible to identify the species at a relatively early stage by using antibodies specific for each species. However, to apply the method, operation for isolating the bacteria is still required. Accordingly, it takes more several days, and it is difficult to obtain sufficient detection sensitivity and selectivity.
More quick detection methods have been studied, lately, as disclosed in Japanese Patent Laid-open Publication Nos. 5-15400(1993) and 6-141899(1994), and as reported in J. Am. Soc. Brew. Chem.: 52(3) 95-99, 1994, there is a method for detecting the lactic acid bacteria, wherein the DNA of lactic acid bacteria is extracted to serve as a sample and an oligonucleotide complementary for the DNA is functionally used as a primer.
On the other hand, lately, it is confirmed that the genus Pectinatus of an obligately anaerobic bacterium exerts a bad influence on the product beer. To detect the genus, a selective separation medium was examined (J. Am. Soc. Brew. Chem.: 52(3) 115-119, 1994), a group for determining the main antigen was identified by using an immunoblotting technique (FEMS. MICROBIOL. LETT.: 67(3) 307-311, 1990), and a method using a fluoroimmunoassay method was disclosed (J. Am. Soc. Brew. Chem.: 51(4) 158-163, 1993). However, satisfactory methods are not established in aspects of the detectable sensitivity, detection times and the like.
The present invention aims to develop a method wherein the genus Pectinatus is more quickly and sensitively detected and identified at the same time.
SUMMARY OF THE INVENTION
The present invention provides a method wherein oligonucleotides specifically hybridizing with 16S ribosomal genes of the genus Pectinatus are prepared. These oligonucleotides are then used for gene amplification by functioning as a primer, and each bacteria is selectively detected.
The oligonucleotides specifically hybridizing Pectinatus cerevisiiphilus have at least one of the following sequence groups:
Further, the oligonucleotides specifically hybridizing Pectinatus frisingensis have at least one of the following sequence groups:
Then, the oligonucleotides, which target a nucleotide sequence coding a 16S ribosomal RNA gene of Pectinatus sp. DSM20764 and are complementary to the sequence n
REFERENCES:
patent: 5484909 (1996-01-01), Nietupski et al.
Schleifer et al., Intl. J. Systematic Bacteriology 40(1):19-27, Jan. 1990.
Doyle et al., J. Industrial Microbiology 15:67-70, 1995.
Asahi Breweries Ltd.
Zitomer Stephanie W.
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