Abstract: Long wavelength, narrow emission bandwidth fluorecein dyes are provided for detecting spacially overlapping target substances. The dyes comprise 4,7-dichlorofluoresceins, and particularly 2',4',5', 7'-tetrachloro-4,7-dichloro-5- (and 6-) carboxyfluoresceins. Methods and kits for using the dyes in DNA analysis are provided.

Abstract: Method and composition for detecting one or more selected polynucleotide regions in a target polynucleotide. In the method, a mixture of sequence-specific probes are reacted with the target polynucleotide under hybridization conditions, and the hybridized probes are treated to selectively modify those probes which are bound to the target polynucleotide in a base-specific manner. The resulting labeled probes include a polymer chain which imparts to each different-sequence probe, a distinctive ratio of charge/translational frictional drag, and a detectable label. The labeled probes are fractionated by electrophoresis in a non-sieving matrix, and the presence of one or more selected sequences in the target polynucleotide are detected according to the observed electrophoretic migration rates of the labeled probes in a non-sieiving medium.

Abstract: Long wavelength, narrow emission bandwidth fluorecein dyes are provided for detecting spacially overlapping target substances. The dyes comprise 4,7-dichlorofluoresceins, and particularly 2',4',5',7'-tetrachloro-4,7-dichloro-5- (and 6-) carboxyfluoresceins. Methods and kits for using the dyes in DNA analysis are provided.

Abstract: Method and composition for detecting one or more selected polynucleotide regions in a target polynucleotide. In the method, a mixture of sequence-specific probes are reacted with the target polynucleotide under hybridization conditions, and the hybridized probes are treated to selectively modify those probes which are bound to the target polynucleotide in a base-specific manner. The resulting labeled probes include a polymer chain which imparts to each different-sequence probe, a distinctive ratio of charge/translational frictional drag, and a detectable label. The labeled probes are fractionated by electrophoresis in a non-sieving matrix, and the presence of one or more selected sequences in the target polynucleotide are detected according to the observed electrophoretic migration rates of the labeled probes in a non-sieving medium.

Abstract: Method and composition for detecting one or more selected polynucleotide regions in a target polynucleotide. In one embodiment of the invention, a plurality of different-sequence probe pairs are added to a target polynucleotide, where each probe pair includes two polynucleotide probe elements which are complementary in sequence to adjacent portions of a selected one of the target sequences in the target polynucleotide. In each probe pair, one of the probe elements contains a non-polynucleotide polymer chain which imparts a distinctive mobility to the associated probe pair, when the elements in the pair are ligated. The other element in the pair contains a detectable reporter label. After the probe pairs have been allowed to hybridize with the target polynucleotide, the hybridized polynucleotides are treated under conditions effective to ligate the end subunits of target-bound probe elements when their end subunits are base-paired with adjacent target bases.

Abstract: Method and composition for detecting one or more selected polynucleotide regions in a target polynucleotide. In the method, a mixture of sequence-specific probes are reacted with the target polynucleotide under hybridization conditions, and the hybridized probes are treated to selectively modify those probes which are bound to the target polynucleotide in a base-specific manner. The resulting labeled probes include a polymer chain which imparts to each different-sequence probe, a distinctive ratio of charge/translational frictional drag, and a detectable label. The labeled probes are fractionated by electrophoresis in a non-sieving matrix, and the presence of one or more selected sequences in the target polynucleotide are detected according to the observed electrophoretic migration rates of the labeled probes in a non-sieving medium.

Abstract: Long wavelength, narrow emission bandwidth fluorecein dyes are provided for detecting spacially overlapping target substances. The dyes comprise 4,7-dichlorofluoresceins, and particularly 2',4',5',7'-tetrachloro-4,7-dichloro-5- (and 6-) carboxyfluoresceins. Methods and kits for using the dyes in DNA analysis are provided.

Abstract: Method and composition for detecting one or more selected polynucleotide regions in a target polynucleotide. In the method, a mixture of sequence-specific probes are reacted with the target polynucleotide under hybridization conditions, and the hybridized probes are treated to selectively modify those probes which are bound to the target polynucleotide in a base-specific manner. The resulting labeled probes include a polymer chain which imparts to each different-sequence probe, a distinctive ratio of charge/translational frictional drag, and a detectable label. The labeled probes are fractionated by electrophoresis in a non-sieving matrix, and the presence of one or more selected sequences in the target polynucleotide are detected according to the observed electrophoretic migration rates of the labeled probes in a non-sieving medium.

Abstract: A method for synthesizing a polynucleotide having a nitrogen atom at the 3'-end is disclosed. The methods include the following steps. A solid support reagent is provided having the structure shown below: ##STR1## where T is an acid-cleavable hydroxyl protecting group, e.g., 4,4'-dimethoxytritil; Q is a linker connecting the nitrogen and oxygen, e.g., n-hexyl; R.sub.1 is a nitrogen substituent, e.g., hydrogen; R.sub.2 through R.sub.4 are separately hydrogen or lower alkyl; Y is an atom which is electronegative with respect to carbon, e.g., oxygen or sulfur; X.sub.1 is an atom which is electronegative with respect to carbon, e.g., sulfone; Z is a bond or spacer arm, e.g., ethylsuccinate; and W is a derivatized solid synthesis support capable of linking to Z, e.g., an amino-dirivitized controlled pore glass. The solid support is treated with acid to remove the acid-cleavable hydroxyl protecting group.

Abstract: Method and kits are provided for detecting one or more target nucleic acids. A first oligonucleotide having a 3' amino group and a second oligonucleotide having a 5' phosphate group are annealed to a contiguous complementary region of a target nucleic acid. Whenever the 3' terminal nucleotides of the first oligonucleotide and the 5' nucleotides of the second oligonucleotide are complementary to the opposing nucleotides on the target nucleic acid, a nucleic acid ligase ligates the first and second oligonucleotides via the formation of a phosphoramidate linkage. The presence of the target nucleic acid is determined by detection of the ligated first and second oligonucleotides.

Abstract: Method and composition for detecting one or more selected polynucleotide regions in a target polynucleotide. In the method, a mixture of sequence-specific probes are reacted with the target polynucleotide under hybridization conditions, and the hybridized probes are treated to selectively modify those probes which are bound to the target polynucleotide in a base-specific manner. The resulting labeled probes include a polymer chain which imparts to each different-sequence probe, a distinctive ratio of charge/translational frictional drag, and a detectable label. The labeled probes are fractionated by electrophoresis in a non-sieving matrix, and the presence of one or more selected sequences in the target polynucleotide are detected according to the observed electrophoretic migration rates of the labeled probes in a non-sieiving medium.

Abstract: The compounds are exemplified by the class of sulfoethyl oxygen-substituted carbamates, such compounds being useful as support reagents for automated polynucleotide synthesis of 3'-nitrogen functionalized polynucleotides. The invention includes, in one aspect, a polynucleotide synthesis reagent having the structure: ##STR1## where T is an acid-cleavable hydroxyl protecting group, e.g., 4,4'-dimethoxytritil; Q is a linker connecting the nitrogen and oxygen, e.g., n-hexyl; R.sub.1 is a nitrogen substituent, e.g., hydrogen; R.sub.2 through R.sub.4 are separately hydrogen or lower alkyl; Y is an atom which is electronegative with respect to carbon, e.g., oxygen or sulfur; X.sub.1 is an atom which is electronegative with respect to carbon, e.g., sulfone; Z is a bond or spacer arm, e.g., ethylsuccinate; and W is a derivatized solid synthesis support capable of linking to Z, e.g., an amino-dirivitized controlled pore glass.

Abstract: Method and composition for detecting one or more selected polynucleotide regions in a target polynucleotide. In one embodiment of the invention, a plurality of different-sequence probe pairs are added to a target polynucleotide, where each probe pair includes two polynucleotide probe elements which are complementary in sequence to adjacent portions of a selected one of the target sequences in the target polynucleotide. In each probe pair, one of the probe elements contains a non-polynucleotide polymer chain which imparts a distinctive mobility to the associated probe pair, when the elements in the pair are ligated. The other element in the pair contains a detectable reporter label. After the probe pairs have been allowed to hybridize with the target polynucleotide, the hybridized polynucleotides are treated under conditions effective to ligate the end subunits of target-bound probe elements when their end subunits are base-paired with adjacent target bases.

Abstract: Method and composition for detecting one or more selected polynucleotide regions in a target polynucleotide. In the method, a mixture of sequence-specific probes are reacted with the target polynucleotide under hybridization conditions, and the hybridized probes are treated to selectively modify those probes which are bound to the target polynucleotide in a base-specific manner. The resulting labeled probes include a polymer chain which imparts to each different-sequence probe, a distinctive ratio of charge/translational frictional drag, and a detectable label. The labeled probes are fractionated by electrophoresis in a non-sieving matrix, and the presence of one or more selected sequences in the target polynucleotide are detected according to the observed electrophoretic migration rates of the labeled probes in a non-sieving medium.

Abstract: A spectrally resolvable set of rhodamine dyes are provided for use in the chain termination method of nucleic acid sequencing. A different rhodamine dye from the group consisting of tetramethylrhodamine, rhodamine X, rhodamine 6G, and rhodamine 110 is attached to the base of each of the dideoxynucleotides used in the sequencing method by way of an alkynylamino linker. Preferably, the labeled dideoxynucleotides are incorporated into the growing DNA chains by Taq DNA polymerase.

Abstract: The compounds of the invention include novel linking agents comprising 2-substituted-3-protected-1,3,2-oxazaphosphacycloalkanes and their phosphoramidite precursors. The compounds of the invention further include conjugates of the above linking agents with oligonucleotides and polymer supports. The compounds of the present invention are useful for linking organic moieties, such as fluorescent or chromogenic dyes, to polymer supports and oligonucleotides, particularly single- and double-stranded DNA and RNA fragments.

Abstract: The invention provides a novel cleavage reagent for hydrolysing base-labile linking groups between a solid phase support and oligonucleotides. The cleavage reagent comprises a lower alkyl alcohol, water, and a non-nuccleophilic hindered alkylamine containing from 3 to 6 carbon atoms in a ratio of about 1:1:1 to about 1:3:1, respectively. An important property of the cleavage reagent is that it preserves the fluorescent characteristics of rhodamine dyes during cleavage, thereby making it possible to completely synthesize rhodamine-labeled oligonucleotides by solid phase methods. Rhodamine phosphoramidites are provided to further enhance the efficiency of synthesizing rhodamine-labeled oligonucleotides by such methods.

Abstract: The compounds of the invention include novel linking agents comprising 2-substituted-3-protected-1,3,2-oxazaphosphacycloalkanes and their phosphoramidite precursors. The compounds of the invention further include conjugates of the above linking agents with oligonucleotides and polymer supports. The compounds of the present invention are useful for linking organic moieties, such as fluorescent or chromogenic dyes, to polymer supports and oligonucleotides, particularly single- and double-stranded DNA and RNA fragments.

Abstract: Long wavelength, narrow emission bandwidth fluorecein dyes are provided for detecting spacially overlapping target substances. The dyes comprise 4,7-dichlorofluoresceins, and particularly 1',2',7',8'-dibenzo-4,7-dichlorofluoresceins. Methods of using the dyes in automated DNA sequencing are described.

Abstract: The invention consists of compounds and methods for the synthesis of oligonucleotides which contain one or more free aliphatic amino groups attached to the sugar moieties of the nucleoside subunits. The synthetic method is versatile and general, permitting amino groups to be selectively placed at any position on oligonucleotides of any composition or length which is attainable by current DNA synthetic methods. Fluorescent dyes or other detectable moieties may be covalently attached to the amino groups to yield the corresponding modified oligonucleotide.