Amoco Corporation

Vysis, Inc.

Other

University of California

Kenneth A Cruickshank

Larry E Morrison

Garfield P Royer

Douglas J Taron

Michael L Bittner

Clarissa F Prorok

Lucy M Stols

Methods, apparatus and compositions are presented for ligating ligands together which bind to a common receptor. One embodiment includes polynucleotide probes having photoreactive functional groups. The probes are capable of assuming substantially contiguous reactive positions on a target polynucleotide placing the photoreactive group in juxtaposition. Activation of the photoreactive functional groups with radiant energy form a probe reaction product in which the probes are bound to each other.

Template-directed photoligation

The invention relates to a method for linking a detectable label to a nucleic acid by (1) providing a nucleic acid bound to a solid support, the nucleic acid having a cytidine base; (2) transaminating the cytidine base with a reactive group to form a covalent linkage between the cytidine base and the reactive group; and (3) linking a detectable label to the reactive group. The invention also includes compositions containing a labeled nucleic acid produced by the methods of the invention immobilized on a solid support, and a kit containing a solid support, a bisulfite, a reactive group, and a detectable label.

Solid phase nucleic acid labeling by transamination

A method for the detection of a target polynucleotide sequence wherein the target polynucleotide sequence is used as a template to direct hybridization of first and second polynucleotide probes to contiguous portions of the target. The hybridized probes are then ligated, preferably by covalent linkage of photoreactive functional groups present on each of the probes, to produce a ligated reaction product. The ligated reaction product is then multiplied with a polymerase such as Q-beta replicase to produce an enzyme reaction product which is then detected. The presence of the enzyme reaction product is indicative of the presence of the target polynucleotide sequence.

Template-directed ligation and amplification assay

This invention discloses methods and compounds for covalently linking xanthine or lower alkyl substituted derivatives of xanthine to DNA and the resulting xanthine or lower alkyl substituted xanthine derivative labeled DNA reagents. Reagents for the in situ detection of a chromosome or a region of a chromosome are disclosed. These reagents include a multiplicity of DNA sequences that are complementary to different portions of the chromosome or chromosome region to be detected. Multiple xanthine or lower alkyl substituted xanthine derivative labels are covalently linked to the DNA sequences. These xanthine or lower alkyl substituted xanthine derivative labeled reagents are contacted under hybridizing conditions with the chromosome or chromosome region of interest. Any binding of the reagent with the chromosome or chromosome region of interest may then be detected by immunological techniques.

Methods and compounds for labeling DNA with xanthine and lower alkyl

Techniques for producing cloned DNA sequences are provided which sequences are complementary to DNA occurring in one selected region of one chromosome of a multi-chromosomal genome, such as the human genome. Such cloned DNA sequences can be labeled and formed into probes by conventional procedures, there are provided methods for making probe compositions which comprise mixed DNA segments derived from such a DNA sequence. An improved DNA sequence transamination procedure is provided utilizing trifluoroacetate chaotrope anions. With high concentrations of low complexity DNA, high levels of transamination are thereby achieved. These segments are covalently bound to fluorophore groups through linking groups that are transaminated preferably chaotropically into the segments.

Production of chromosome region specific DNA sequences and transamination

Methods, apparatus and compositions are presented for ligating ligands together which bind to a common target. One embodiment includes polynucleotide probes having photoreactive functional groups. The probes are capable of assuming substantially contiguous reactive positions on a target polynucleotide placing the photoreactive groups in juxtaposition. Activation of the photoreactive functional groups with radiant energy form a probe reaction product in which the probes are bound to each other.

This invention discloses methods and compounds for covalently linking nitro-, dinitro- and trinitrophenyl to DNA and the resulting nitro-, dinitro-, and trinitrophenyl labeled DNA reagents. Reagents for the in situ detection of a chromosome or a region of a chromosome are disclosed. These reagents include a multiplicity of DNA sequences that are complementary to different portions of the chromosome or chromosome region to be detected. Multiple nitro-, dinitro- or trinitrophenyl labels are covalently linked to the DNA sequences by means of an amino acid linking group. These labeled reagents are contacted under hybridizing conditions with the chromosome or chromosome region of interest. Any binding of the reagent with the chromosome or chromosome region of interest may then be detected by immunological techniques.

Labeling reagents useful for the chemical attachment of nitrophenyl

The present invention includes nucleotide compositions having linking groups and the use and manner of making the nucleotide compositions. The nucleotide compositions are used to attach nonisotopic label moieties to nucleic acid probes.

Nucleotide compositions with linking groups

This invention encompasses diagnostic reagents comprising a polynucleic acid probe having a specific binding sequence and having one or more cytidines outside of the specific binding sequence. The polynucleic acid probe is bound to an amino functionalized solid support by a bisulfite mediated transamination between a cytidine and an amino group on the solid support. When the specific binding sequence contains cytidine, the cytidine is replaced with 5-methylcytidine which has essentially the same binding capacity and yet does not participate in transamination reactions. The invention encompasses compositions of the polynucleic acid probe as well as test kits including the above identified reagent and assays utilizing the above reagent. This invention is useful in detecting viruses, fungi and bacteria in test samples such as body fluids and food samples as well as detecting DNA or RNA sequences in mammalian cells.

Growing community of inventors

Naperville, IL, United States of America

Kenneth A Cruickshank