Microarray slide for detecting toxic algae, barcodes of DNA, hybridisation solution and method of detecting toxic algae
Inventors
MEDLIN, Linda • MONTRESOR, Marina • GRANELI, Edna • REGUERA, Beatriz • RAINE, Robin • EDVARDSEN, Bente • Lewis, Jane
Assignees
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Publication Number
US-10294533-B2
Publication Date
2019-05-21
Expiration Date
Abstract
An array system is provided including a microarray slide configured to detect simultaneously a plurality of organisms in a sample, wherein the microarray slide includes nucleic acid probes having fragments of 18S or 28S RNA sequence unique to each organism or taxonomical group thereof. The present microarray slide is useful for detecting different species of toxic algae.
Core Innovation
The invention provides an array system comprising a microarray slide configured to detect simultaneously a plurality of organisms in a sample. The microarray slide comprises nucleic acid probes having fragments of 18S or 28S RNA sequence that are unique to each organism or to a taxonomical group thereof, and the probes include a poly-T tail or an alternative linker or a spacer element.
The probe sets cover a hierarchical taxonomic structure. Some nucleic acid probes are specific for detection of species of each organism to be detected, while other nucleic acid probes are specific for detection of at least one higher-level taxon to which each organism belongs, including taxonomical phylum, taxonomical class, an intermediate taxonomical level clade between class and genus, taxonomical genus, and taxonomical strain where both toxic and non-toxic strains of the same species exist.
The slide comprises positive and negative controls as well as probe sequences defined by SEQ ID NO ranges. In the described approach, the system is used for rapid detection, identification, and quantification of toxic algae in marine samples using nucleic acid probes with fluorescent labeling, and the rationale presented includes hierarchical calls with thresholding and reduced reliance on mouse bioassays.
Claims Coverage
Two independent claims are directed to an array system using a microarray slide with hierarchical nucleic acid probes targeting unique 18S/28S RNA fragments, including probe-tail/linker features and structured probe sets spanning controls and taxonomic levels. Across both independent claims, the inventive features are substantially the same, with differences in the specificity of control/group requirements as written, and the claims collectively cover species through strain for organisms and toxic/non-toxic strain discrimination for a species where both exist.
Simultaneous multi-organism detection with 18S/28S RNA unique fragments
A microarray slide configured to detect simultaneously a plurality of organisms in a sample, wherein the microarray slide comprises nucleic acid probes having fragments of 18S or 28S RNA sequence unique to each organism or taxonomical group thereof.
Probe tail or alternative linker or spacer element
Each of the nucleic acid probes has a poly-T tail or an alternative linker or a spacer element.
Hierarchical taxonomic probe coverage from species to strain
Some of the nucleic acid probes are specific for detection of the species of each organism to be detected and other nucleic acid probes are specific for detection of at least one higher-level taxon to which each of the organisms to be detected belongs, including taxonomical phylum, taxonomical class, an intermediate taxonomical level clade between class and genus, taxonomical genus, and taxonomical strain where both toxic and non-toxic strains of the same species exist.
Positive and negative controls with defined SEQ ID probe sets
The microarray slide comprises at least one probe selected from the group consisting of positive and negative controls, with probe sequences defined by SEQ ID NO ranges.
The independent claims define an array system that combines multi-organism simultaneous detection using microarray probes based on unique 18S/28S RNA sequence fragments, probe structural features, and a hierarchical set of taxon-selective probes spanning phylum through strain with toxic versus non-toxic strain detection, together with positive and negative controls and SEQ ID defined probe groups.
Stated Advantages
Rapid detection, identification, and quantification of toxic algae in marine samples.
High-confidence hierarchical calls with thresholding.
Ability to infer cell concentrations via calibration curves.
Reduced reliance on mouse bioassays.
Documented Applications
Detection, identification, and quantification of toxic algae in marine samples.
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