- Substractive single label comparative hybridization
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Provided are methods of determining differences between nucleic acids in a test sample and a reference sample. In certain embodiments the methods are used for detecting and mapping chromosomal or genetic abnormalities associated with various diseases or with predisposition to various diseases, or to detecting the phenomena of large scale copy number variants. In particular, provided are advanced methods of performing array-based comparative hybridization that allow reproducibility between samples and enhanced sensitivity by using the same detectable label for both test sample and reference sample nucleic acids. Invention methods are useful for the detection or diagnosis of particular disease conditions such as cancer, and detecting predisposition to cancer based on detection of chromosomal or genetic abnormalities and gene expression level. Invention methods are also useful for the detection or diagnosis of hereditary genetic disorders or predisposition thereto, especially in prenatal samples. Moreover, invention methods are also useful for the detection or diagnosis of de novo genetic aberrations associated with post-natal developmental abnormalities.
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- Activatable probes and methods for in vivo gene detection
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Probes for detecting a target polynucleotide are provided. One aspect provides a molecular beacon probe set wherein the donor molecular beacon comprises a quantum dot and an acceptor molecular beacon comprises at least one reporter. The probes optionally comprise a protein transduction domain, targeting signal, or a combination thereof. Methods for detecting target polynucleotides using the disclosed probes are also provided.
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- Oligonucleotides comprising a molecular switch
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This invention relates to oligonucleotides comprising a molecular switch which may exist in an “open” or “closed” position. The molecular switch portion of the probe is particularly sensitive to the identity of sequences complementary to the molecular switch. Oligonucleotides containing a molecular switch are applicable to all kinds of hybridization processes. Due to the sensitivity of the switch domain of the oligonucleotide, probes containing a molecular switch are particularly useful in the identification of single point mismatches. More specifically, a portion, but not all, of the oligonucleotide becomes unbound from a mismatched target. The invention further relates to methods of using said oligonucleotides for research reagents, and clinical diagnostics. An exemplary oligonucleotide comprises a first hybridizable domain, a second bridging block domain, and a third binding domain.
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- Reactive derivatives of sulforhodamine 101 with enhanced hydrolytic stability
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The invention describes reactive dyes having an alkyl spacer attached via a sulfonamide bond to a sulforhodamine 101 fluorophore, and a variety of useful conjugates prepared therefrom. The increased length of the covalent linkage due to the alkyl spacer results in dye-conjugates having a number of surprisingly advantageous properties relative to previous sulforhodamine 101-labeled conjugates, including enhanced solubility and increased fluorescence. The reactive dyes of the present invention are more stable than the known compound sulforhodamine 101 sulfonyl chloride. Novel reactive dyes are described for selective modification of groups other than amines, including thiols and photoreactive derivatives.
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- Dual resonance energy transfer nucleic acid probes
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Dual nucleic acid probes with resonance energy transfer moieties are provided. In particular, fluorescent or luminescent resonance energy transfer moieties are provided on hairpin stem-loop molecular beacon probes that hybridize sufficiently near each other on a subject nucleic acid, e.g. mRNA, to generate an observable interaction. The invention also provides lanthanide chelate luminescent resonance energy transfer moieties on linear and stem-loop probes that hybridize sufficiently near each other on a subject nucleic acid to generate an observable interaction. The invention thereby provides detectable signals for rapid, specific and sensitive hybridization determination in vivo. The probes are used in methods of detection of nucleic acid target hybridization for the identification and quantification of tissue and cell-specific gene expression levels, including response to external stimuli, such as drug candidates, and genetic variations associated with disease, such as cancer.
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- Nucleic acid amplification oligonucleotides with molecular energy transfer labels and methods based thereon
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The present invention provides labeled nucleic acid amplification oligonucleotides, which can be linear or hairpin primers or blocking oligonucleotides. The oligonucleotides of the invention are labeled with donor and/or acceptor moieties of molecular energy transfer pairs. The moieties can be fluorophores, such that fluorescent energy emitted by the donor is absorbed by the acceptor. The acceptor may be a fluorophore that fluoresces at a wavelength different from the donor moiety, or it may be a quencher. The oligonucleotides of the invention are configured so that a donor moiety and an acceptor moiety are incorporated into the amplification product. The invention also provides methods and kits for directly detecting amplification products employing the nucleic acid amplification primers. When labeled linear primers are used, treatment with exonuclease or by using specific temperature eliminates the need for separation of unincorporated primers. This "closed-tube" format greatly reduces the possibility of carryover contamination with amplification products, provides for high throughput of samples, and may be totally automated.
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