Abstract: The invention provides methods and compositions for amplifying selected polynucleotides, especially selected subsets of restriction fragments. Generally, methods of the invention are implemented by ligating adaptors containing at least one promoter sequence to such fragments under conditions that promote the formation of closed single stranded or double stranded structures, which are capable of serving as cyclical templates for transcription.

Abstract: The invention provides methods, kits and materials for determinining simultaneously signature sequences of a population of tagged polynucleotides. Tags comprise at least two parts: a hybridization tag and a correlation tag. Size ladders of polynucleotide fragments are generated from the population of tagged polynucleotides that contain a plurality of size classes. After the size classes are separated, hybridization tags of the separated fragments are copied and labeled according to the identity of one or more bases at the ends of the fragments. In a preferred embodiment, the labeled tags are specifically hybridized to a plurality of random microarrays of tag complements. Signals generated at hybridization sites of different random microarrays are correlated by sequencing of the unique correlation tag. Signature sequences are determined by signals generated at hybridization sites having the same correlation tag on each of the plurality of random microarrays.

Abstract: A method and compositions are provided for analyzing whole genomes, or representations thereof, to determine associations between traits and genotypes. Sets of hybridization probes (referred to herein as “isostringency probes”) are provided that are complementary to sites uniformly spaced throughout unique sequence regions a genome, or target polynucleotide, and that are designed for facile isolation of subsets that form perfectly matched duplexes with the genome or target polynucleotide being analyzed. The nucleotide sequences of the isostringency probes are selected to ensure that the probes have substantially identical duplex stabilities. In accordance with tie method of the invention, representations of a genome are attached to solid phase supports and are used to capture isostringency probes forming perfectly matched duplexes. The captured probes are then released and applied to an array of complementary sequences for detection.

Abstract: The invention provides a method for constructing a high resolution physical map of a polynucleotide. In accordance with the invention, nucleotide sequences are determined at the ends of restriction fragments produced by a plurality of digestions with a plurality of combinations of restriction endonucleases so that a pair of nucleotide sequences is obtained for each restriction fragment. A physical map of the polynucleotide is constructed by ordering the pairs of sequences by matching the identical sequences among the pairs.

Abstract: The invention provides a method for constructing a high resolution physical map of a polynucleotide. In accordance with the invention, nucleotide sequences are determined at the ends of restriction fragments produced by a plurality of digestions with a plurality of combinations of restriction endonucleases so that a pair of nucleotide sequences is obtained for each restriction fragment. A physical map of the polynucleotide is constructed by ordering the pairs of sequences by matching the identical sequences among the pairs.

Abstract: Compositions and methods are disclosed for detecting multiple target analytes, particularly polynucleotide target analytes. In accordance with one aspect of the invention, a template-dependent extension reaction is performed to generate detection probes, such that each detection probe has (i) at least one molecular tag attached by a cleavable linkage and (ii) either a capture moiety or a cleavage-inducing moiety attached. The template-dependent extension reaction may be carried out directly on a polynucleotide analyte to generate molecular tags, wherein the polynucleotide analyte serves as a template in the template-dependent extension reaction, or it may be carried out indirectly on an oligonucleotide label that, in turn, is attached to a binding moiety specific for an analyte of interest. In either case, a plurality of molecular tags are generated, after which they are separated and identified to determine the presence or absence or the quantity of the target analytes in a sample.

Abstract: The present invention includes, in one aspect, a kit comprising one or more sets of oligonucleotide probes whose perfectly matched duplexes with complementary sequences have similar or substantially the same free energy of duplex formation.

Abstract: The invention provides a method of tracking, identifying, and/or sorting classes or subpopulations of molecules by the use of oligonucleotide tags. Oligonucleotide tags of the invention comprise oligonucleotides selected from a minimally cross-hybridizing set. Preferably, such oligonucleotides each consist of a plurality of subunits 3 to 9 nucleotides in length. A subunit of a minimally cross-hybridizing set forms a duplex or triplex having two or more mismatches with the complement of any other subunit of the same set. The number of oligonucleotide tags available in a particular embodiment depends on the number of subunits per tag and on the length of the subunit. An important aspect of the invention is the use of the oligonucleotide tags for sorting polynucleotides by specifically hybridizing tags attached to the polynucleotides to their complements on solid phase supports.

Abstract: The invention provides a method of tracking, identifying, and/or sorting classes or subpopulations of molecules by the use of oligonucleotide tags. Oligonucleotide tags of the invention comprise oligonucleotides selected from a minimally cross-hybridizing set. Preferably, such oligonucleotides each consist of a plurality of subunits 3 to 9 nucleotides in length. A subunit of a minimally cross-hybridizing set forms a duplex or triplex having two or more mismatches with the complement of any other subunit of the same set The number of oligonucleotide tags available in a particular embodiment depends on the number of subunits per tag and on the length of the subunit. An important aspect of the invention is the use of the oligonucleotide tags for sorting polynucleotides by specifically hybridizing tags attached to the polynucleotides to their complements on solid phase supports.

Abstract: The invention provides a method of tracking, identifying, and/or sorting classes or subpopulations of molecules by the use of oligonucleotide tags. Oligonucleotide tags of the invention comprise oligonucleotides selected from a minimally cross-hybridizing set. Preferably, such oligonucleotides each consist of a plurality of subunits 3 to 9 nucleotides in length. A subunit of a minimally cross-hybridizing set forms a duplex or triplex having two or more mismatches with the complement of any other subunit of the same set. The number of oligonucleotide tags available in a particular embodiment depends on the number of subunits per tag and on the length of the subunit.An important aspect of the invention is the use of the oligonucleotide tags for sorting polynucleotides by specifically hybridizing tags attached to the polynucleotides to their complements on solid phase supports.

Abstract: The invention provides a method and materials for analyzing the frequency of sequences in a population of polynucleotides, such as a cDNA library. A population of restriction fragments is formed which is inserted into vectors which allow segments to be removed from each end of the inserted fragments. The segments from each restriction fragment are ligated together to form a pair of segments which serves as a tag for the restriction fragment, and the polynucleotide from which the fragment is derived. Pairs of segments are excised from the vectors and ligated to form concatemers which are cloned and sequenced. A tabulation of the sequences of pairs provides a frequency distribution of sequences in the population.

Abstract: The invention provides a method for constructing a high resolution physical map of a polynucleotide. In accordance with the invention, nucleotide sequences are determined at the ends of restriction fragments produced by a plurality of digestions with a plurality of combinations of restriction endonucleases so that a pair of nucleotide sequences is obtained for each restriction fragment. A physical map of the polynucleotide is constructed by ordering the pairs of sequences by matching the identical sequences among the pairs.

Abstract: Disclosed are oligonucleotides probes which are useful for determining a sequence of nucleotides in a target ploynucleotide. In one embodiment, the probe is of the formula: HO-(3')(B)j(5')-OP(.dbd.O-)NH-(B)k-Bt*, wherein B is a nucleotide or an analog thereof; j is in range of from 1 to 12; is in the range of from 0 to 12, such that the sum of j and k is less than or equal to 12; and Bt* is a labeled chain-terminating moiety. In s second embodiment, the probe has a formula selected from the group consisting of (1) OP(.dbd.O)(O--)O--(5.dbd.)(B)sRRRR(B)wBt*, and (2) OP(.dbd.O)(O--)(5')(B)sRRRR(B)w(3')OP(.dbd.O)(O--)O, wherein: is a deoxyribonucleotide or an analog thereof; R is a ribonucleotide; s is in the range of from 1 to 8; w is in the range of from 0 to 8, such that the sum of j and k is less than or equal to 8; and Bt* is a labeled chain-terminating moiety.

Abstract: The invention provides a method of tracking, identifying, and/or sorting classes or subpopulations of molecules by the use of oligonucleotide tags. Oligonucleotide tags of the invention comprise oligonucleotides selected from a minimally cross-hybridizing set. Preferably, such oligonucleotides each consist of a plurality of subunits 3 to 9 nucleotides in length. A subunit of a minimally cross-hybridizing set forms a duplex or triplex having two or more mismatches with the complement of any other subunit of the same set. The number of oligonucleotide tags available in a particular embodiment depends on the number of subunits per tag and on the length of the subunit. An important aspect of the invention is the use of the oligonucleotide tags for sorting polynucleotides by specifically hybridizing tags attached to the polynucleotides to their complements on solid phase supports.

Abstract: Method and compositions are provided for analyzing nucleic acid sequences based on repeated cycles of duplex extension along a single stranded template. Preferably, such extension starts from a duplex formed between an initializing oligonucleotide and the template. The initializing oligonucleotide is extended in an initial extension cycle by ligating an oligonucleotide probe to its end to form an extended duplex. The extended duplex is then repeatedly extended by subsequent cycles of ligation. During each cycle, the identity of one or more nucleotides in the template is determined by a label on, or associated with, a successfully ligated oligonucleotide probe. Preferably, the oligonucleotide probe has a blocking moiety, e.g. a chain-terminating nucleotide, in a terminal position so that only a single extension of the extended duplex takes place in a single cycle. The duplex is further extended in subsequent cycles by removing the blocking moiety and regenerating an extendable terminus.

Abstract: A method is provided for sequencing nucleic acids without the need to separate similarly sized DNAs or RNAs by gel electrophoresis. The method relies on the separate hybridization of multiple mixed oligonucleotide probes to a target sequence. The mixed oligonucleotide probes comprise sequences of fixed and non-fixed bases corresponding to every possible permutation of fixed and non-fixed bases less than or equal to the length of the probes. For each probe, the hybridizations provide the number of times the probe's particular sequence of fixed bases appears in the target sequence. The target sequence is then mathematically reconstructed from this data and a knowledge of the probe sequences.