SYBR Real Time PCR Kit (SYBR Premix Ex Taq Kit)

Introduction

SYBR Premix Ex Taq Kit is used for intercalator based real time PCR using SYBR®Green.

Rapid reaction mixtures are based on a 2× concentration premixed with SYBR®Green I at a concentration appropriate for real time monitoring.

A combination of Ex Taq™ HS, a hot start PCR enzyme that uses an anti-Taq antibody, and a buffer optimized for real time PCR allows high amplification efficiency and high detection sensitivity in real time PCR. The 2× premixed reagent also contains Tli RNaseH, a heat-resistant RNase H, which minimizes inhibition of PCR due to residual mRNA when using cDNA as template. This product is suitable for high-speed PCR and allows accurate assay and detection of targets, making it possible to conduct real time PCR analyses with good reproducibility and high reliability.

Components

1.  DNA Extraction, 5mL,2 tubes

2.  SYBR Premix Ex Taq (2×Conc.) (5units,1.0 mL) 

DNA Preparation

-Blood

1. Take 500μL anticoagulant blood to a 1.5 ml clean centrifuge tube, add 1 ml of ddH₂O, shake for 30s .

2. centrifuge at 8,000 rpm for 5 min, discard the supernatant.

3. Repeat washing until no red precipitate.

4. Add 1mL physiological saline, shake 15s, centrifuge at 10,000 rpm for 5 min, discard the supernatant.

5. Add 50μL DNA extraction and mix with precipitation.

6. 100 ℃ 10 min. Centrifuged at 13,000 rpm for 3 min.

7. take the supernatant 4μL for PCR reactions.

-Animal tissue, food or feed

1.  Take about 30 mg homogenized samples, and put

into a 1.5 mL clean centrifuge tube, add 100μL DNA extraction and mix well (the DNA extraction must dissolve at room temperature and fully mixed before use).

2.  Water bath at 56 ℃ for 30 min, then at 100 ℃ for 10 min.

3.  Centrifuge at 13,000 rpm for 5 min.

4.  Transfer the supernatant 5μL for PCR reactions.

Real Time PCR Reaction

1.  Take out qPCR MIX from the kit, melt it at room temperature and oscillate it, then mix it with10000 rpm centrifugal 10 s;

2.  Each of the 25μL system, the upstream and downstream primers were 0.5-1 μL, respectively, mixed and amplificated.

3.  Example: IQTM5, ABI PRISM7500 and other instruments using thin walled tubes: circulation conditions: 94℃ for 2 minutes, 

     then 94℃for 30s, 60℃ for 30 s,72℃ for 30 s, 30 cycles. The user should set the corresponding amplification conditions 

      according to their own primers, and the above conditions are for reference only.

Specification

   50 preps,100 preps,200 preps,250 preps.