Organic compounds -- part of the class 532-570 series – Organic compounds – Carbohydrates or derivatives
Reexamination Certificate
2000-06-30
2003-01-28
Marschel, Ardin H. (Department: 1631)
Organic compounds -- part of the class 532-570 series
Organic compounds
Carbohydrates or derivatives
C435S006120, C436S501000, C536S023100, C536S024310, C536S024320, C536S025300, C702S020000
Reexamination Certificate
active
06512105
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The inventions described and claimed herein relate to probes and assays based on the use of genetic material such as RNA. More particularly, the inventions relate to the design and construction of nucleic acid probes and hybridization of such probes to genetic material of target non-viral organisms in assays for detection and/or quantitation thereof in test samples of, e.g., sputum, urine, blood and tissue sections, food, soil and water.
2. Introduction
Two single strands of nucleic acid, comprised of nucleotides, may associate (“hybridize”) to form a double helical structure in which the two polynucleotide chains running in opposite directions are held together by hydrogen bonds (a weak form of chemical bond) between pairs of matched, centrally located compounds known as “bases.” Generally, in the double helical structure of nucleic acids, for example, the base adenine (A) is hydrogen bonded to the base thymine (T) or uracil (U) while the base guanine (G) is hydrogen bonded to the base cytosine (C). At any point along the chain, therefore, one may find the base pairs AT or AU, TA or UA, GC, or CG. One may also find AG and GU base pairs in addition to the traditional (“canonical”) base pairs. Assuming that a first single strand of nucleic acid is sufficiently complementary to a second and that the two are brought together under conditions which will promote their hybridization, double stranded nucleic acid will result. Under appropriate conditions, DNA/DNA, RNA/DNA, or RNA/RNA hybrids may be formed.
Broadly, there are two basic nucleic acid hybridization procedures. In one, known as “in solution” hybridization, both a “probe” nucleic acid sequence and nucleic acid molecules from a test sample are free in solution. In the other method, the sample nucleic acid is usually immobilized on a solid support and the probe sequence is free in solution.
A probe may be a single strand nucleic acid sequence which is complementary in some particular degree to the nucleic acid sequences sought to be detected (“target sequences”). It may also be labelled. A background description of the use of nucleic acid hybridization as a procedure for the detection of particular nucleic acid sequences is described in U.S. application Ser. No. 456,729, entitled “Method for Detection, Identification and Quantitation of Non-Viral Organisms,” filed Jan. 10, 1983 (Kohne I, now issued as U.S. Pat. No. 4,851,330), and U.S. application Ser. No. 655,365, entitled “Method For Detecting, Identifying and Quantitating organisms and Viruses,” filed Sep. 4, 1984 (Kohne II, now issued as U.S. Pat. No. 5,288,611), both of which are incorporated by reference, together with all other applications cited herein.
Also described in those applications are methods for determining the presence of RNA-containing organisms in a sample which might contain such organisms, comprising the steps of bringing together any nucleic acids from a sample and a probe comprising nucleic acid molecules which are shorter than the rRNA subunit sequence from which it was derived and which are sufficiently complementary to hybridize to the rRNA of one or more non-viral organisms or groups of non-viral organisms, incubating the mixture under specified hybridization conditions, and assaying the resulting mixture for hybridization of the probe and any test sample rRNA. The invention is described to include using a probe which detects only rRNA subunit subsequences which are the same or sufficiently similar in particular organisms or groups of organisms and is said to detect the presence or absence of any one or more of those particular organisms in a sample, even in the presence of many non-related organisms.
We have discovered and describe herein a novel method and means for designing and constructing DNA probes for use in detecting unique rRNA sequences in an assay for the detection and/or quantitation of any group of non-viral organisms. Some of the inventive probes herein may be used to detect and/or quantify a single species or strain of non-viral organism and others may be used to detect and/or quantify members of an entire genus or desired phylogenetic grouping.
SUMMARY OF THE INVENTION
In a method of probe preparation and use, a single strand deoxyoligonucleotide of particular sequence and defined length is used in a hybridization assay to determine the presence or amount of rRNA from particular target non-viral organisms to distinguish them from their known closest phylogenetic neighbors. Probe sequences which are specific, respectively, for 16S rRNA variable subsequences of
Mycobacterium avium, Mycobacterium intracellulare
and the
Mycobacterium tuberculosis
-complex bacteria, and which do not cross react with nucleic acids from each other, or any other bacterial species or respiratory infectious agent, under proper stringency, are described and claimed. A probe specific to three 23S rRNA variable region subsequences from the
Mycobacterium tuberculosis
-complex bacteria is also described and claimed, as are rRNA variable region probes useful in hybridization assays for the genus Mycobacterium (16S 23S rRNA specific),
Mycoplasma pneumoniae
(5S and 16S rRNA-specific),
Chlamydia trachomatis
(16S and 23S rRNA specific),
Enterobacter cloacae
(23S rRNA specific),
Escherichia coli
(16S rRNA specific), Legionella (16S and 23s rRNA specific), Salmonella (16S and 23S rRNA specific), Enterococci (16S rRNA specific),
Neisseria gonorrhoeae
(16S rRNA specific), Campylobacter (16S rRNA specific),
Proteus mirabilis
(23S rRNA specific), Pseudomonas (23S rRNA specific), fungi (18S and 28S rRNA specific), and bacteria (16S and 23S rRNA specific).
In one embodiment of the assay method, a test sample is first subjected to conditions which release rRNA from any non-viral organisms present in that sample. rRNA is single stranded and therefore available for hybridization with sufficiently complementary genetic material once so released. Contact between a probe, which can be labelled, and the rRNA target may be carried out in solution under conditions which promote hybridization between the two strands. The reaction mixture is then assayed for the presence of hybridized probe. Numerous advantages of the present method for the detection of non-viral organisms over prior art techniques, including accuracy, simplicity, economy and speed will appear more fully from the detailed description which follows.
REFERENCES:
patent: 3755086 (1973-08-01), Heimer
patent: 3930956 (1976-01-01), Juni
patent: 4033143 (1977-07-01), Juni
patent: 4228238 (1980-10-01), Swanson
patent: 4237224 (1980-12-01), Cohen et al.
patent: 4275149 (1981-06-01), Litman et al.
patent: 4302204 (1981-11-01), Wahl et al.
patent: 4358535 (1982-11-01), Falkow et al.
patent: 4394443 (1983-07-01), Weissman et al.
patent: 4416988 (1983-11-01), Rubin
patent: 4480040 (1984-10-01), Owens et al.
patent: 4677054 (1987-06-01), White et al.
patent: 4689295 (1987-08-01), Taber et al.
patent: 4711955 (1987-12-01), Ward et al.
patent: 4717653 (1988-01-01), Webster
patent: 4851330 (1989-07-01), Kohne
patent: 5087558 (1992-02-01), Webster
patent: 5089386 (1992-02-01), Stackebrandt et al.
patent: 5091519 (1992-02-01), Cruickshank
patent: 5217862 (1993-06-01), Barns et al.
patent: 5288611 (1994-02-01), Kohne
patent: 5348854 (1994-09-01), Webster
patent: 5447848 (1995-09-01), Barns et al.
patent: 5840488 (1998-11-01), Hogan
patent: 5851767 (1998-12-01), Stanbridge et al.
patent: 6150517 (2000-11-01), Hogan et al.
patent: 3138784 (1985-02-01), None
patent: 0155359 (1982-09-01), None
patent: 0079139 (1982-10-01), None
patent: 0120658 (1984-03-01), None
patent: 0133671 (1984-07-01), None
patent: 0232085 (1987-08-01), None
patent: 0245129 (1987-11-01), None
patent: 0250662 (1988-01-01), None
patent: 0277237 (1988-08-01), None
patent: 0155360 (1988-12-01), None
patent: 8301073 (1983-03-01), None
patent: 8401174 (1984-03-01), None
patent: 8402721 (1984-07-01), None
patent: 8803957 (1988-06-01), None
Palmer et al., Chlamydial Infections—Proceedings of the Sixth Int
Hogan James John
Kop Jo Ann
McDonough Sherrol Hoffa
Smith Richard Dana
Cappellari Charles B.
Fisher Carlos A.
Gen-Probe Incorporated
Heber Sheldon O.
Marschel Ardin H.
LandOfFree
Methods for making oligonucleotide probes for the detection... does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Methods for making oligonucleotide probes for the detection..., we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Methods for making oligonucleotide probes for the detection... will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-3042672