qPCR 2x Blue Master Mix_BLUE

Product description

This product is an alternative to qPCR 2x Blue Master Mix enriched with a dye qPCR Visible Blue Mark from Top-Bio (Cat. No. B129). The dye does not interfere with qPCR, but facilitates visualization of Master Mixes presence in multi-well PCR plates. The Mix is used mostly for qPCR with DNA-specific probes such as TaqMan, Molecular beacons, FRET and others.

Rapid preparation (2x concentrated)

• The qPCR 2x Blue Master Mix_BLUE possesses all components 2x concentrated (optimized reaction buffer, nucleotides, Taq DNA polymerase and anti-Taq monoclonal antibody). The samples are prepared simply by mixing an aliquot of the Mix with oligonucleotide primers, template DNA, H₂O (included) and selected DNA probes. This facilitates rapid preparation of the PCRs.

Hot start

• The product contains monoclonal antibody anti-Taq, which binds to Taq DNA polymerase and thus inactivates its enzymatic activity. After the first denaturation cycle, the antibody is irreversibly inactivated, and Taq DNA polymerase regains enzymatic activity. This decreases the formation of nonspecific DNA amplicons.

Sensitive detection

• This product is optimized for the sensitive detection of DNA fragments amplified during qPCR from genomic DNA or cDNA obtained by reverse transcription.

 Rapid setup

• qPCR 2x Blue Master Mix_BLUE is especially useful for routine analyses of large numbers of DNA samples. To 0.5 ml of the Master Mix in the original tube, primers (e.g. 40 µl forward and 40 µl reverse), PCR H₂O and fluorescent probes are added and mixed; the "armed Mix" can be stored at -20 ± 5^oC. Immediately before PCR, the Mix is thawed, and each 24 µl aliquot is mixed with 1 µl of the tested DNA template.

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 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 Blue Master Mix (Top-Bio, Cat. No. P521) was supplemented with qPCR Visible Blue Mark (blue lines; final dilution 1:200) or not (green lines), SYBR Green 1, 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 Blue Master Mix_BLUE (this amount is sufficient e.g. for 40 reactions, 25 µl each).
• 1 tube with 1.5 ml PCR H₂O.

Storage

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

Composition

• The Mix is 2x concentrated: 150 mM Tris-HCl, pH 8. 8 (at 25oC), 40 mM (NH4)2SO4, 0.02% Tween 20, 5 mM MgCl2, 400 µM dATP, 400 µM dCTP, 400 µM dGTP, 400 µM dTTP, Taq DNA polymerase (50 U/ml), monoclonal antibody anti-Taq (38 nM), qPCR Visible Blue Mark, stabilizers and additives.

Purity and quality control

• Each batch of qPCR 2x Blue Master Mix_BLUE is tested for amplification of a single copy gene in genomic DNA.