The present invention provides a DNA polymerase including the sequence of SEQ ID NO. 1 or a sequence which is at least 70% identical thereto, but wherein the aspartic acid residue at position 18 of SEQ ID NO. 1, or the equivalent aspartic acid residue in other sequences, has been replaced by a non-negatively charged amino acid residue. It further provides DNA polymerases comprising the amino acid sequences of SEQ ID NO. 2, 11 and 12 and variants thereof. The present invention also provides nucleic acids encoding the DNA polymerases, a method of producing said DNA polymerases, and compositions, expression vectors and host cells or viruses comprising said DNA polymerases. The present invention also provides uses of said DNA polymerases in nucleotide polymerisation, amplification, and sequencing reactions.

An apparatus for preparing nucleic acid sequences using an enzyme, including a reactor, a plurality of nucleotide material bottles, a deblocking material bottle, and a liquid delivering device. The reactor includes a reaction substrate having a pretreated surface. Each of the nucleotide material bottles is adapted to contain a first reaction solution, and the first reaction solution includes a reaction enzyme and a nucleotide having a terminal protecting group. The deblocking material bottle is adapted to contain a deblocking solution. The liquid delivering device is connected to the reactor, the nucleotide material bottles and the deblocking material bottle. The reaction enzyme is adapted to dispose the nucleotide having the terminal protecting group on the pretreated surface. The reactor has an operating temperature of 45° C.-105° C. A method for preparing nucleic acid sequences using an enzyme of the invention is provided.

Methods of amplifying an RNA template according to aspects of the present disclosure include: providing a composition including a recombinant thermostable DNA polymerase including SEQ ID NO:1, or a variant thereof having at least 99% identity to SEQ ID NO:1; and a reaction buffer, the reaction buffer including 10-30 mM Tris-HCl, pH 8.5-9.0; 20-40 mM KCl; 5-15 mM (NH.sub.4).sub.2SO.sub.4; 1.5-3.5 mM Mn.sup.2+; 0.01-0.20% (w/v) gelatin and/or serum albumin; 0.05-0.15% (w/v) of a nonionic detergent; with the proviso that no more than 0.1 mM of Mg.sup.2+ is present in the composition; and, optionally, 0.01-0.1 mM of a chelating agent is present in the composition.

The invention includes a mutant Taq polymerase or biologically active fragment thereof, which can significantly extend and amplify a target sequence in comparison to wild type Taq polymerase, where the extension mixture includes asymmetrical cyanine dye, including SYBR Green I. The mutant Taq polymerase, or a biologically active fragment thereof, has one or more substitutions differing from the wild type as shown in Table I.

Provided are compositions comprising recombinant DNA polymerases that include amino acid substitutions, insertions, deletions, and/or exogenous features that confer modified properties upon the polymerase for enhanced single molecule sequencing. Such properties include increased resistance to photodamage, and can also include enhanced metal ion coordination, reduced exonuclease activity, reduced reaction rates at one or more steps of the polymerase kinetic cycle, decreased branching fraction, altered cofactor selectivity, increased yield, increased thermostability, increased accuracy, increased speed, increased readlength, and the like. Also provided are nucleic acids which encode the polymerases with the aforementioned phenotypes, as well as methods of using such polymerases to make a DNA or to sequence a DNA template.

Modified polypeptides having polymerase activity.

This disclosure relates to novel compounds for use in various compositions, kits and methods, including, for example, use in polymerase storage buffers and in nucleic acid synthesis or amplification reactions such as a polymerase chain reaction (PCR). Methods for preparing the novel compounds are also described.

The present invention provides improved DNA polymerases, in particular, type A DNA polymerases, that may be better suited for applications in recombinant DNA technologies. Among other things, the present invention provides modified DNA polymerases derived from directed evolution experiments designed to select mutations that confer advantageous phenotypes under conditions used in industrial or research applications.

The invention relates to mutant DNA polymerases of the Pol theta subfamily capable of performing non-templated nucleic acid extension, or of a functional fragment of such a polymerase, methods of producing these mutant DNA polymerases, kits and methods of using these mutant DNA polymerases.

The present invention is related to a method for adding one or more L-nucleotides to the 3′end of a first L-nucleic acid, wherein the method comprises the step of reacting the one or more L-nucleotides with the first L-nucleic acid in the presence of a protein comprising a mutant enzymatic activity exhibiting moiety, wherein the enzymatic activity is capable of adding one or more L-nucleotides to the 3′ end of the first L-nucleic acid, wherein the mutant enzymatic activity exhibiting moiety comprises an amino acid sequence, wherein the amino acids of the amino acid sequence are D-amino acids, wherein the mutant enzymatic activity exhibiting moiety is a variant of an enzymatic activity exhibiting moiety, wherein the enzymatic activity exhibiting moiety consists of an amino acid sequence according to SEQ ID NO: 15 and wherein the amino acids of the amino acid sequence according to SEQ ID NO: 15 are D-amino acids, wherein the amino acid sequence of the mutant enzymatic activity exhibiting moiety differs from the amino acid sequence of the enzymatic activity exhibiting moiety consisting of an amino acid sequence according to SEQ ID NO: 15 at least at one amino acid position, preferably at three amino acid positions, and/or wherein the amino acid sequence of the mutant enzymatic activity exhibiting moiety is a truncated form of an amino acid sequence according to SEQ ID NO: 15, and wherein the amino acid sequence of the mutant enzymatic activity exhibiting moiety is different from an amino acid sequence according to any of SEQ ID NOs 15 to 22 and 51.