qPCR 2x SYBR Master Mix_BLUE

Product description

Description

qPCR 2x SYBR Master Mix_BLUE is dedicated to qPCR quantification of DNA amplicons with fluorescent DNA dye SYBR Green I; it also contains a blue dye, which does not interfere with qPCR and facilitates visualization of Master Mixes in multi-well plates.

SYBR Green I

• The Mix contains intercalating DNA dye SYBR Green I, which after binding to double-stranded (ds)DNA, becomes strongly fluorescent with maximal excitation at 497 nm (blue light) and emission at 520 nm (green light). Because the fluorescence of unbound SYBR Green I is very low, enhanced fluorescence during qPCR corresponds to an increase in dsDNA amplicons produced during PCR.

Blue dye

• The Mix also contains a qPCR Visible Blue Mark dye from Top-Bio (Cat. No. B129) that enhances the visibility of real-time PCR reactions for more accurate pipetting, plate loading, and reaction tracking.

Hot start

• The Mix contains an anti-Taq DNA polymerase monoclonal antibody, which inactivates the enzymatic activity of the enzyme. After the first denaturation cycle, the antibody is irreversibly inactivated, and Taq DNA polymerase regains enzymatic activity.

Rapid samples preparation

• All components of the qPCR 2x SYBR Master Mix_BLUE are 2x concentrated, facilitating rapid PCR sample preparation. The samples are prepared by mixing an aliquot of the Mix with oligonucleotide primers, template DNA and H₂O (included).
• qPCR 2x SYBR Master Mix_BLUE is especially useful for routine analyses of large DNA samples. For example, to 0.5 ml of the Master Mix in the original tube, primers (e.g. 40 µl forward and 40 µl reverse) and PCR H₂O are added and mixed; the "armed" Mix can be stored at -20 ± 5^o Immediately before use, the "armed" Mix is thawed and aliquoted into reaction wells. After addition of the DNA template, a qPCR is performed.

Figure 1. qPCR Visible Blue Mark facilitates pipetting into multi-well plates. Wells of the 384-well plate were pipetted with 10 µl aliquotes of qPCR 2x Blue Master Mix (columns 3-8) or 10 µl aliquotes of qPCR 2x Blue Master Mix_BLUE containing 100-fold diluted qPCR Visible Blue Mark (columns 1, 2, 9-14, 23, 24). In columns 16-22, qPCR 2xSYBR Master Mix_BLUE was pipetted only in rows A, C, E, G, I, K, M and O. In column 15, all wells were left blank.

Figure 2. qPCR Visible Blue Mark does not affect the qPCR performance. qPCR 2x SYBR Master Mix (Top-Bio, Cat. No. P551) was supplemented with qPCR Visible Blue Mark (blue lines; final dilution 1:200) or not (green lines), DNA template at five 10-fold dilutions, and the corresponding primers. No DNA template control (NTC) was also included. The samples in triplicates were analyzed by a real-time PCR cycler.

Technical data

Components and packaging

• 1 tube with 0.5 ml qPCR 2x SYBR Master Mix_BLUE (for 40 reactions, 25 µl each).

• 1 tube with 1.5 ml PCR Ultra H₂O.

Composition       

• qPCR 2x SYBR Master Mix_BLUE contains: 20 mM Tris-HCl, pH 8. 8 (at 25^oC), 100 mM KCl, 0.2% Triton X-100, 3 mM MgCl₂, 400 µM dATP, 400 µM dCTP, 400 µM dGTP, 400 µM dTTP, 50 U/ml Taq DNA polymerase, monoclonal antibody anti-Taq (38 nM), SYBR Green I, qPCR Visible Blue Mark, stabilizers and additives.

Storage

• At temperature -20^oC ± 5°C. Material can be repeatedly defrosted.

Purity and quality control              

• The quality of DNA polymerase is verified by SDS PAGE; only one band of 94 kDa is observed in Coomassie blue-stained gel. Material is free of nucleases.
• Each batch of qPCR 2x SYBR Master Mix_BLUE is tested for amplification of a single copy gene in genomic DNA.