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BioMaster RT-PCR SYBR Blue (2×)


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BioMaster RT-PCR SYBR Blue (2×) reagent kit includes 2× buffer for RT-PCR with SYBR containing all the necessary components (except for DNA template and primers), intercalating dye SYBR Green I, BioMaster-mix and DEPC-treated water. The kit is designed for one-step reverse transcription and real-time polymerase chain reaction (qRT-PCR).
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-PCR with SYBR is optimized for both efficient RT and real-time PCR. It contains additives and enhancers that enable efficient RT-PCR of complicated and GC-rich templates. Blue hue of the reaction solution provided by the inert dye allows control when using multi-well plates.
 
Product composition
 
Cat. #
2× buffer for RT-PCR with SYBR BioMaster-mix  
DMSO
DEPC-treated water Number of reactions
(50 μl each)
RM04-40 2 × 0.5 ml 1 × 80 µl 0.5 ml 2 × 0.5 ml 40
RM04-200 4 × 1.25 ml 2 × 200 µl 0.5 ml 3 × 1.8 ml 200
2× buffer for RT-PCR with SYBR contains:
100 mM Tris-НCl (рН 8.3 at 25 °С), 150 mM KCl, 0.6 mM each deoxynucleoside triphosphate, 6 mM MgCl2, 8 mM DTT, enzyme stabilizers and enhancers, fluorescent dye SYBR Green I and inert dye.
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 and HS-Taq DNA polymerase.
 
Applications:
  • Gene expression analysis;
  • One-step real-time RT-PCR.
 
Reaction mix features
  • 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-PCR SYBR Blue (2×) at room temperature for 2 days and/or multiple thawing-freezing cycles do not affect RT-PCR efficacy);
  • The mix contains fluorescent dye SYBR Green I, which allows real-time monitoring of PCR.
 
SYBR Green I
SYBR Green I is a fluorescent intercalating dye for quantitative and qualitative detection of PCR products during real-time PCR. SYBR Green I provides easy and economical way for detection and quantitative assessment of PCR products during real-time PCR without a need of using specific fluorescent probes. During amplification, SYBR Green I dye penetrates into the minor groove of DNA products and emits stronger fluorescent signal than the unbound dye. Absorption and emission maxima of SYBR Green I are 494 nm and 521 nm, respectively, which enables its usage with any real-time PCR platform existing to date.
 
Inert dye
The inert dye included in 2× buffer for RT-PCR with SYBR does not reduce PCR efficiency; it facilitates monitoring of multi-well plate pipetting. Absorption maximum of the blue dye is 615 nm.
 
Benefits of use
  • The mix is colored for easy pipetting
  • High specificity
  • High sensitivity (1pg – 1 µg of RNA)
  • 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)
 
Limits of use
The presence of the fluorescent dye renders usage of fluorescent probes.

Storage and transportation: at -20 °С; not more than 30 thawing-freezing cycles.

Storage terms: 9 months (under proper storage and transportation conditions).