Abstract: This invention concerns non-standard nucleotides that can form non-standard Watson-Crick nucleobase pairs having geometries similar to the geometries of standard nucleotide pairs, but that are joined by a non-standard hydrogen bonding schemes. Disclosed are processes that yield oligonucleotides that are semi-complementary to a standard oligonucleotide, where the region of semi-complementarity pairs one or more standard nucleotides with a non-standard nucleotide, or vice versa. Duplexes formed from two semi-complementary oligonucleotides are also inventions disclosed.

Abstract: This invention provides processes to assemble many (greater than 20) partially overlapping single stranded DNA molecules (fragments) having preselected sequences, followed by extension of those strands that hybridize at terminal overlap regions, and ligation of the extend products, creating a double-stranded DNA assembly. These processes use non-standard nucleotides carrying heterocyclic nucleobase analogs that implement non-standard hydrogen bonding patterns; these allow controlled annealing of the single stranded fragments via Watson-Crick rules, with less or no interference from a range of non-Watson Crick interactions, hairpin formations, or off-target hybridization displayed by standard nucleobases. This process includes an optional conversion step that replaces non-standard nucleobase pairs with standard nucleobase pairs, generating large synthetic DNA (LS-DNA) molecules containing only natural nucleotides.

Abstract: This invention combines artificially expanded genetic information systems (AEGIS) with self-avoiding molecular recognition systems (SAMRS), in processes that involve template-directed primer extension in highly multiplexed form in mixtures containing large numbers of primers. This process yields extension products, or in its PCR format, amplicons, that have AEGIS tags that can be cleanly captured in highly complex mixtures.

Abstract: This invention concerns self-avoiding molecular recognition systems (SAMRS), compositions that bind to natural DNA and RNA, but do not bind to compositions at sites that incorporate other SAMRS components, and processes dependent on them. Their utility is shown by discoveries that DNA polymerases accept these compositions as primers and templates, where standard triphosphates are added to primers containing SAMRS components, and added opposite to SAMRS components in the template. A critical mass of data are provided in 16 examples to provide first-generation heuristic rules to permit design of SAMRS sequences can be used as primers and templates that are accepted by DNA polymerases. The presently preferred primers are at least 12 nucleotide units in length, and more preferably between 15 and 30 nucleotides in length. Also preferred are chimeric primers that have standard nucleotides in their 5?-segments, and SAMRS nucleotides in their 3?-segments, and in multiplexed priming.

Abstract: The disclosed invention teaches processes to amplify oligonucleotides by contacting templates and primers with DNA polymerases and triphosphates of non-standard nucleotides, which form nucleobase pairs fitting the standard Watson-Crick geometry, but joined by hydrogen bonding patterns different from those that join standard A:T and G:C pairs. Thus, this invention relates to nucleotide analogs and their derivatives that, when incorporated into DNA and RNA, expand the number of replicatable nucleotides beyond the four found in standard DNA and RNA. The invention further relates to polymerases that incorporate those non-standard nucleotide analogs into oligonucleotide products using the corresponding triphosphate derivatives, and more specifically, polymerases and non-standard nucleoside triphosphates that support the polymerase chain reaction (PCR), including PCR where the products contain more than one non-standard nucleotide unit.

Abstract: This invention enables processes for extracting from a mixture of oligonucleotide molecules individuals that bind to a preselected target (aptamers) or catalyze a preselected reaction (xNAzymes) that contain one or more non-standard nucleotides. These pair with their complements in a Watson-Crick geometry with a pattern of hydrogen bonds different from that pairing adenine and thymine, and guanine and cytosine. The processes comprise (a) obtaining this mixture containing non-standard nucleotides within preselected regions, (b) contacting the mixture with a preselected target or one or more reactants for the reaction whose catalysis is desired, (c) separating oligonucleotides having a greater affinity to the target or catalytic effectiveness from the remainder that have less affinity or catalytic effectiveness, and (d) amplifying the separated oligonucleotides.

Abstract: This invention relates to nucleoside, nucleotide, and oligonucleotide analogs that incorporate non-standard nucleobase analogs, defined to be those that present a pattern of hydrogen bonds to a paired nucleobase analog in a complementary strand that is different from the pattern presented by adenine, guanine, cytosine, and thymine. Most specifically, this invention discloses and claims processes for amplifying nucleic acid analogs containing non-standard nucleobases using polymerase chain reactions, and enzymes that perform this amplification.

Abstract: This invention relates to the field of nucleic acid chemistry, more specifically to the field of compositions of matter that comprise triphosphates of modified 2?-deoxynucleosides and oligonucleotides that are formed when these are appended to the 3?-end of a primer, wherein said modifications comprise NH2 moiety attached to their 3?-hydroxyl group and a fluorescent species in a form of a tag affixed to the nucleobase via a linker that can be cleaved. Such compositions and their associated processes enable and improve the sequencing of oligonucleotides using a strategy of cyclic reversible termination, as outlined in U.S. Pat. No. 6,664,079. Most specifically, the invention concerns compositions of matter that are 5?-triphosphates of ribo- and 2?-deoxyribonucleosides carrying detectable tags and oligonucleotides that might be derived from them.

Abstract: This invention relates to the field of nucleic acid chemistry, more specifically to compositions of matter that are nucleic acid analogs, and processes that use them. Still more specifically, these compositions comprise two fragments of DNA-like molecules, each having one or more ends modified to carry a reactive group, where the reactive group on one fragment can form a transient covalent bond with the reactive group on the other under conditions of dynamic equilibrium to form a composite, where the composite can then bind to a target oligonucleotide, such as a DNA or RNA molecule. Most specifically, once the transient covalent bond forms, the composite serves as a primer for a template-directed polymerization using a DNA polymerase, an RNA polymerase, or a reverse transcriptase. Once incorporated, the epimerization causes the base pair to be destabilized, the duplex containing the epimerized nucleoside to likewise be destabilized, and the double strand to then disassociate.

Abstract: This invention relates to nucleoside, nucleotide, and oligonucleotide analogs that incorporate non-standard nucleobase analogs, defined to be those that present a pattern of hydrogen bonds to a paired nucleobase analog in a complementary strand that is different from the pattern presented by adenine, guanine, cytosine, and thymine. The invention is specifically concerned with compositions of matter that present the donor-donor-acceptor, donor-acceptor-donor, and acceptor-donor-donor non-standard hydrogen bonding patterns on pyrimidine analogs, where nucleoside analogs bearing these pyrimidine analogs do not epimerize as easily as those known in the art. The heterocycles on these nucleoside analogs are diaminopyridines and aminopyridones that have electron withdrawing groups attached to the position analogous to the 5-position of the ring in standard pyrimidines, including nitro, cyano, and carboxylic acid derivatives.

Abstract: Processes are disclosed that use 3?-reversibly terminated nucleoside triphosphates to analyze DNA for purposes other than sequencing using cyclic reversible termination. These processes are based on the unexpected ability of terminal transferase to accept these triphosphates as substrates, the unexpected ability of polymerases to add reversibly and irreversibly terminated triphosphates in competition with each other, the development of cleavage conditions to remove the terminating group rapidly, in high yield, and without substantial damage to the terminated oligonucleotide product, and the ability of reversibly terminated primer extension products to capture groups. The presently preferred embodiments of the disclosed processes use a triphosphate having its 3?-OH group blocked as a 3?-ONH2 group, which can be removed in buffered NaNO2 and use variants of Taq DNA polymerase, including one that has a replacement (L616A).

Abstract: A process is provided to generate fluorescent molecules in the presence of target nucleic acids, but not in the absence of that target. Two probes, one bearing moiety A and the other bearing moiety B, bind to the target in a way that brings A and B together. A photon then converts A into A*, which can react with B to form a new species Z that is fluorescent. If A* does not encounter B, then A* reverts to form A. This allows the probe to have another opportunity to be activated should it be later bound near B. In one embodiment, a photoenolization creates A* as a diene; this reacts with a dienophile B in a Diels-Alder reaction. The linker Z may cause the linked probes to bind less tightly to the target, allowing the target to generate many fluorescent products, or be read through by a polymerase.

Abstract: The disclosed invention teaches processes to amplify oligonucleotides by contacting templates and primers with DNA polymerases and triphosphates of non-standard nucleotides, which form nucleobase pairs fitting the standard Watson-Crick geometry, but joined by hydrogen bonding patterns different from those that join standard A:T and G:C pairs. Thus, this invention relates to nucleotide analogs and their derivatives that, when incorporated into DNA and RNA, expand the number of replicatable nucleotides beyond the four found in standard DNA and RNA. The invention further relates to polymerases that incorporate those non-standard nucleotide analogs into oligonucleotide products using the corresponding triphosphate derivatives, and more specifically, polymerases and non-standard nucleoside triphosphates that support the polymerase chain reaction (PCR), including PCR where the products contain more than one non-standard nucleotide unit.

Abstract: Specific sequences of DNA are often detected by a process that comprises a step where the sequence to be detected (the “analyte”) binds to give a duplex with a DNA molecule or analog that is complementary in the Watson-Crick sense to some portion of the analyte in an aqueous “assay environment” that may contain buffer, salt, and/or detergent. Such purely aqueous systems cannot be exposed indefinitely to the environment, however, as the water in the system will evaporate. Further, such systems often support the growth of bacteria and other organisms, destroying their effectiveness. This invention provides for compositions of matter and processes that use them that comprise assay mixtures containing more than 40% formamide. This mixture remains a liquid at equilibrium with water in environments normally inhabited by humans. This invention also provides for mixtures containing formamides that include detergents.

Abstract: This invention relates to nucleoside, nucleotide, and oligonucleotide analogs that incorporate non-standard nucleobase analogs, those that present a pattern of hydrogen bonds to a paired nucleobase analog in a complementary strand that is different from the pattern presented by adenine, guanine, cytosine, and thymine. Most specifically, this invention discloses and claims processes for amplifying nucleic acid analogs containing non-standard nucleobases using polymerase chain reactions, and combinations of non-standard nucleobases, analogs of standard nucleotides, and enzymes that perform this amplification.

Abstract: This invention provides compositions of matter that, when incorporated into an oligonucleotide, present to a complementary strand in a Watson-Crick pairing geometry a pattern of hydrogen bonds that is different from the pattern presented by adenine, guanine, cytosine, and thymine. Most specifically, this invention discloses and claims compositions of matter that present the same hydrogen bonding patterns as the isocytidine and isoguanosine nucleobases, but do not have unfavorable tautomeric forms, do not become disassociated from their sugar, and do not make major groove interactions, as much, as easily, or as strongly as isocytidine and isoguanosine.

Abstract: This invention relates to the field of nucleic acid chemistry, more specifically to the field of compositions and processes that are nucleic acid analogs. More specifically, it relates to purine analogs that contain three rings, where the third ring bridges the exocyclic substituent at position 6 to position 7, using the purine numbering system. Still more specifically it relates to analogs having this structure that are able to form nucleobase pairs having the Watson-Crick geometry with a pyrimidine or pyrimidine analog, where the nucleobase pair is joined by hydrogen bonding patterns that either present a standard hydrogen bonding pattern, or a non-standard hydrogen bonding pattern. Most specifically, it to nucleoside analogs that are analogs of isoguanosine, but where the 5-6 ring system of the purine ring in isoguanosine is fused to another five- or six-membered ring, where the fused ring joins the exocyclic amino group with an atom that is, by analogy, at position 7 of the isoguanine ring system.

Abstract: This application claims processes and compositions that enable discovery of single nucleotide polymorphisms (SNPs) and other sequence variation that follows two essentially identical sequences, one a reference, the other a target, as well as SNPs discovered using these processes and compositions. The inventive process comprises preparation of four sets of primers, “T-extendable”, “A-extendable”, “C-extendable”, and “G-extendable”. These primers, when templated on a reference genome, add (respectively) T, A, C, and G to their 3?-ends. The invention also comprises a step where these primer sets are separately bound to complementary sequences on target DNA and, once bound, prime extension reactions using target DNA as the template. If the target DNA directs incorporation of the same nucleotide as the reference DNA, then the T-, A-, C-, and G-extendable primers are extended (respectively) by T, A, C, and G.

Abstract: This invention relates to the field of nucleic acid chemistry, more specifically to the field of compositions and processes that can bind to nucleic acids, including compositions and processes that by doing so have use in the capture, detection and sequencing of nucleic acids, and most specifically to nucleoside analogs that incorporate nucleobase analogs that are able to bind to different natural nucleobases with nearly equal affinity, and therefore display a degree of ambiguity, or universality, in their binding pattern. Such nucleobase analogs, when incorporated into oligonucleotides, allow the oligonucleotides to bind with nearly equal affinity and/or melting temperature, to a complementary strand, template the synthesis of oligonucleotide from primers that have a distribution of sequences, and support the sequencing of oligonucleotides. Thus, these have use in human diagnostics, especially when polymorphisms are present in a population.

Abstract: This invention relates to the field of nucleic acid chemistry, more specifically to the field of compositions and processes that are nucleic acid analogs. More specifically, this invention relates to compositions that allow the sequencing of oligonucleotides by synthesis, and processes for sequencing by synthesis that exploit these compositions. Most specifically, the instant invention discloses compositions of matter that are 5?-triphosphates of ribo- and 2?-deoxyribonucleosides wherein the 3?-OH group is replaced by a 3?-ONHR group in the alpha configuration, wherein R is either a H or CH3 group. Also disclosed are these triphosphates where the nucleobase carries, via a linker, a reporter groups, such as a fluorescent species that can be used in single- or multi-copy DNA sequencing, or a tag that can be visualized by ultramicroscopy. Also disclosed are processes that use these compositions to do sequencing by synthesis.