Methods for simplifying microbial nucleic acids by chemical modification of cytosines
Inventors
Millar, Douglas Spencer • Miklos, George L. Gabor
Assignees
Interested in licensing this patent?
MTEC can help explore whether this patent might be available for licensing for your application.
Publication Number
US-8598088-B2
Publication Date
2013-12-03
Expiration Date
Abstract
A method for detecting a microorganism comprising reducing the complexity of a microbial genome or microbial nucleic acid by generating a simplified form of the microbial genome or microbial nucleic acid in which substantially all of the positions naturally occupied by cytosines are occupied by uracil or thymine; and assaying for a microbial specific nucleic acid in the simplified form of the microbial genome or microbial nucleic acid, wherein presence of the microbial specific nucleic acid is indicative of the microorganism.
Core Innovation
The invention relates to detecting a microorganism by reducing the complexity of a microbial genome or a microbial nucleic acid. A derivative form is generated in which substantially all of the positions naturally occupied by cytosines are occupied by uracils in the derivative form by treating the microbial genome or the microbial nucleic acid with an agent selected from bisulphite, acetate and citrate, wherein the agent modifies cytosine to uracil.
The derivative form is then used to obtain a nucleic acid being specific for a microorganism having the microbial genome or the microbial nucleic acid. Detection of the microbial-specific nucleic acid in the derivative form is indicative of the presence of a microorganism having the microbial genome or the microbial nucleic acid.
Dependent claim set refinements specify amplification of the derivative form so that uracil positions become thymine after amplification. The amplification is performed by polymerase chain reaction (PCR), isothermal amplification, or signal amplification, and further refinements include specifying the agent as sodium bisulphite and converting microbial RNA to DNA prior to performing the derivative-generation and detection workflow.
Claims Coverage
The provided set includes one independent claim. The independent claim defines a complete detection method with two core inventive features: genomic or nucleic-acid complexity reduction via cytosine-to-uracil derivative generation, and detection of a microorganism-specific nucleic acid in the derivative form to indicate presence of the microorganism. Dependent claims add amplification-related transformation and specific agent and preprocessing selections.
Cytosine-to-uracil derivative generation to reduce microbial nucleic-acid complexity
A method for detecting a microorganism comprising reducing the complexity of a microbial genome or a microbial nucleic acid by generating a derivative form in which substantially all positions naturally occupied by cytosines are occupied by uracils by treating the microbial genome or the microbial nucleic acid with an agent selected from bisulphite, acetate and citrate, wherein the agent modifies cytosine to uracil.
Derivative-form microorganism-specific nucleic-acid detection as presence indication
The derivative form contains a nucleic acid being specific for a microorganism having the microbial genome or the microbial nucleic acid, and detecting the microbial-specific nucleic acid in the derivative form is indicative of the presence of a microorganism having the microbial genome or the microbial nucleic acid.
Overall, the claim coverage centers on generating a derivative form that substantially replaces cytosines with uracils using bisulphite, acetate or citrate, and then detecting a microorganism-specific nucleic acid within that derivative form to indicate the microorganism’s presence. Dependent claims further specify uracil-to-thymine transformation after amplification, selectable amplification modalities, sodium bisulphite, and microbial RNA-to-DNA conversion.
Stated Advantages
Not explicitly described in patent.
Documented Applications
Not explicitly described in patent.
Interested in licensing this patent?