Application

The High Fidelity PCR Master is used for:PCR up to 5kbRT-PCR up to 5kb

DNA molecular weight marker II has been used to estimate the nucleic acid concentrations corresponding to the intensities of fluorescence obtained.

DNA Molecular Weight Marker II provides accurate sizing of fragments over a broad range of sizes. The fragments have 5′-protruding ends and can be labeled with radioactive nucleotides (e.g. [³²P]-dTTP or [³²P]-dGTP) by standard filling-in reactions. End-labeling reactions can be performed with a radioactive or nonradioactive dideoxynucleotide (e.g., DIG-11-ddUTP) and Terminal Transferase.DNA Molecular Weight Marker for agarose gels; simplifies accurate molecular weight determination of double-stranded DNA fragments generated by: Restriction digests PCR RT-PCR

DNA Molecular Weight Marker II provides accurate sizing of fragments over a broad range of sizes. It is used as a DNA Molecular Weight Marker for agarose gels and simplifies accurate molecular weight determination of double-stranded DNA fragments generated by: Restriction digests PCR RT-PCR

Features and Benefits

The High Fidelity PCR Master reduces the setup time and can be immediately used without thawing.

General description

The High Fidelity PCR Master is a ready-to-use, double-concentrated master mix that contains all the reagents (PCR-Grade dNTPs, MgCl₂, and an optimized reaction buffer) required to perform PCR (except template and primer). PCR grade water is also supplied.The special enzyme blend is composed of two thermostable polymerases: Taq DNA Polymerase and a thermostable polymerase with proofreading activity. The 5′→3′ polymerase activity of Taq DNA Polymerase and the 3′→5′ exonuclease of the proofreading polymerase combine for improved performance. It gives PCR products with high yield, high fidelity, and high specificity from episomal and genomic DNA. Due to the inherent 3′→5′ exonuclease activity of the proofreading polymerase, the High Fidelity PCR Master results in approximately three times the fidelity and approximately twice the yield of Taq DNA Polymerase alone. The product is ideal for blunt end cloning or T/A cloning. It can amplify longer fragments (up to 5kb) with high fidelity and also minimize contamination with the help of two pipetting steps.

DNA Molecular Weight Marker II is a ready-to-use solution, 250µg/ml, in TE buffer (Tris-HCl, 1mM EDTA, pH 8.0). The bacteriophage λ DNA is fragmented in a restriction digestion with Hind III endonuclease. The digestion reaction results in eight double-stranded DNA fragments with the following base pair lengths (1 base pair = 660 Daltons): 125, 564, 2027, 2322, 4361, 6557, 9416, and 23130. The fragments have 5′-protruding ends and can be labeled with radioactive nucleotides (e.g., [³²P]-dTTP or [³²P]-dGTP) by standard filling-in reactions. End-labeling reactions can be performed with a radioactive or nonradioactive dideoxynucleotide (e.g., DIG-11-ddUTP) and Terminal Transferase. The product is also available as DNA Molecular Weight Marker II, Digoxigenin-labeled.

Other Notes

Fragment lengths are derived from computer analysis of the DNA sequence. Depending on the size range of the marker, the smallest fragments will only be visible on overloaded gels.For life science research only. Not for use in diagnostic procedures.

Preparation Note

Storage conditions (working solution): 2 to 8 °COnce thawed store at 2 to 8 °C.

Sequence

The bacteriophage lambda (λ) DNA is fragmented in a restriction digestion with Hind III endonuclease. The digestion reaction results in eight double-stranded DNA fragments with the following base pair lengths (1 base pair = 660 Daltons): 125, 564, 2027, 2322, 4361, 6557, 9416, and 23130.Note: Fragment lengths are derived from computer analysis of the DNA sequence. Depending on the size range of the marker, the smallest fragments will only be visible on overloaded gels.

Storage and Stability

Store at -15–-25 °C. (unopened vial)

Unit Definition

50 µg = 1A₂₆₀ unit