A polynucleotide sequencing method includes a wash step that employs a composition including a polymerase. The composition may also include a plurality of nucleotides. The composition may be configured to prevent the polymerase from incorporating one of the plurality of nucleotides into a copy polynucleotide strand. The composition may be substantially free of Mg.sup.2+.

The present invention relates to the field of molecular biology detection, more particularly to a method for fluorescent quantitative PCR. The method includes: 1) mixing an upstream and downstream primer pair, a fluorescent probe, and a PCR amplification reagent; and 2) carrying out the fluorescent quantitative PCR, where the fluorescent probe has two quenching groups, in which a first quenching group is located at a 3′ end and a second quenching group is labeled on a T base and is 10-15 nt apart from the first quenching group. Using the method for fluorescent quantitative PCR, background signals in the fluorescent quantitative PCR can be reduced; furthermore, the sensitivity of PCR can be improved, the occurrence of false negatives in detection can be reduced, and the amplification efficiency of the fluorescent quantitative PCR can also be improved.

Methods and compositions for protecting DNA from light-induced damage and other modifications that occur during DNA sequencing using fluorescent dyes are disclosed.

Methods and compositions for protecting DNA from light-induced damage and other modifications that occur during DNA sequencing using fluorescent dyes are disclosed.

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.

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.

Provided herein are systems and methods for high-resolution mapping of DNA polymerase fidelity using nucleotide imbalances and next-generation sequencing.

Provided herein are systems and methods for high-resolution mapping of DNA polymerase fidelity using nucleotide imbalances and next-generation sequencing.

The present invention relates to the use of a single-strand DNA binding protein (SSB) which exhibits at least 50% of its maximum ssDNA binding capability in the presence of 500 mM of sodium ions, to dehybridize a DNA molecule or to prevent hybridisation of a complementary ssDNA, wherein the SSB comprises the amino acid sequence of SEQ ID NO:1 or an amino acid sequence which is at least 75% identical to SEQ ID NO:1, or a functional fragment thereof, and wherein the DNA molecule or ssDNA is present in or exposed to a solution containing one or more of the following (i) at least 350 mM of sodium ions; (ii) at least 50 mM of potassium ions; (iii) at least 150 mM of magnesium ions; or (iv) at least 200 mM of calcium ions.

The present invention relates to the use of a single-strand DNA binding protein (SSB) which exhibits at least 50% of its maximum ssDNA binding capability in the presence of 500 mM of sodium ions, to dehybridize a DNA molecule or to prevent hybridisation of a complementary ssDNA, wherein the SSB comprises the amino acid sequence of SEQ ID NO:1 or an amino acid sequence which is at least 75% identical to SEQ ID NO:1, or a functional fragment thereof, and wherein the DNA molecule or ssDNA is present in or exposed to a solution containing one or more of the following (i) at least 350 mM of sodium ions; (ii) at least 50 mM of potassium ions; (iii) at least 150 mM of magnesium ions; or (iv) at least 200 mM of calcium ions.