BioMaster RT-qPCR (2×)

Application
  • Gene expression analysis;
  • One-step RT-PCR.

Basic description

BioMaster RT-qPCR (2×) reagent kit includes 2× buffer for RT-qPCR containing all components (except for DNA template and primers), 25× BioMaster-mix and DEPC-treated water. The kit is designed for one-step reverse transcription and real-time polymerase chain reaction (RT-qPCR) with fluorescent probes.


25× BioMaster-mix contains the optimal ratio of M-MuLV –RH to HS-Taq DNA polymerase for both reactions.

M-MuLV –RH is a genetically modified Moloney Murine Leukemia Virus reverse transcriptase (M-MuLV). The enzyme exerts RNA- and DNA-dependent polymerase activity but lacks RNase H activity. M-MuLV –RH reverse transcriptase exhibits improved thermal stability and is active at high temperatures (up to 50°С).

HS-Taq DNA polymerase is a recombinant Taq DNA Polymerase inactivated by specific monoclonal antibodies. The enzyme is inactive at temperatures up to 70 °C, it is activated at the first PCR cycle during a short 5-min incubation at 95 °C. Recombinant HS-Taq DNA polymerase catalyzes 5’→3’ synthesis of DNA and possesses 5’→3’ exonuclease activity of the native Taq DNA polymerase from Thermus aquaticus. Recombinant HS-Taq DNA polymerase is ideal for conventional PCR of the templates of up to 5 kbp.

2× buffer for RT-qPCR is optimized for both efficient RT and PCR.
 
Product composition

 
Cat. #
2× buffer for RT-qPCR 25× BioMaster-mix  
DMSO
DEPC-treated water Number of reactions
(50 μl each)
RM03-40 2 × 0.5 ml 1 × 80 µl 0.5 ml 2 × 0.5 ml 40
RM03-200 4 × 1.25 ml  2 × 200 µl 0.5 ml 3 × 1.8 ml 200
               
 
2× buffer for RT-qPCR contains:
100 mM Tris-НCl (рН 8.3 at 25 °С), 150 mM KCl, 0.6 mM of each deoxynucleoside triphosphate, 6 mM MgCl2, 10 mM TCEP, enzyme stabilizers and enhancers.

25× BioMaster-mix contains:
50 mM Tris-НCl (рН 8.0 at 25 °С), 100 mM NaCl, 1 mM EDTA, 5 mM DTT, 50 % (v/v) glycerol and 0.1 % (v/v) NP-40, M-MuLV –RH reverse transcriptase, highly-processive HS-Taq DNA polymerase and inhibitor of RNases.

Limits of use
Not recommended to use for amplicons of ˃ 5 kbp

Areas of use

  • Gene Expression Analysis
  • RT-PCR one- and two-step

Reaction mixture properties

  • The reaction mix is optimized for the specific and effective performance of M-MuLV –RH reverse transcriptase and HS-Taq DNA polymerase;
  • The mix contains substances that allow long-term storage (storage of BioMaster RT-qPCR (2×) at room temperature for 2 days does not affect RT-PCR efficacy) and multiple thawing-freezing cycles;
  • Does not contain dyes, which makes the reaction mix multi-purpose.

Benefits of using

  • Increased specificity
  • High sensitivity
  • Easy and convenient in use
  • Low pipetting error and low risk of cross-contamination
  • Standardized conditions of the same-type reactions (reduced pipetting error during mixing PCR components in a series of experiments)
  • PCR products can be further subjected to TA cloning due to deoxyadenosine overhangs at the ends of amplified DNA fragments.

Storage

at +4 ° C – 1 week; at -20 ° С - 1 year; not more than 30 thawing-freezing cycles.

Transportation

at 0 - +4 ° C, it is allowed to transport at room temperature up to 8 days.

Product passport

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