Ordinary PCR

Taq DNA Polymerase 说明书

Product number:PC 3015

Product Introduction:Taq DNA polymerase is a highly persistent 5 '→ 3' DNA polymerase that lacks 3 '→ 5' exonuclease activity. It is a single polypeptide chain with a molecular weight of about 95kDa. Taq DNA polymerase was originally isolated from the thermophilic eubacterial Thermus aquaticus BM, which lacks the Taq I restriction enzyme. The enzyme has been cloned into Escherichia coli.

Product specifications:1 ml

Save condition:-30 ~ -15°C storage, ≤ 0°C transport

Scope of application:Taq DNA polymerase has high purity and quality standards and can be used for: PCR, RT-PCR, qPCR, and other primer extension reactions, such as sequencing and labeling. Taq DNA polymerase has no proofreading or hot-start function and is optimally active at 75°C and pH 9.

2 ×Tap Master Mix Instruction Manual

Product number:PC 3021

Product Introduction:This product is a high-purity heat-resistant DNA polymerase, which is expressed by Escherichia coli cloned with Thermus aquaticus DNAPolymerase gene and purified by multiple steps. It does not contain endonuclease, exonuclease and bacterial DNA. The product includes Taq DNA Polymerase, dNTPs, and an optimized buffer system that allows amplification with the addition of primers and templates, reducing pipetting operations and improving throughput and reproducibility of results. This product provides a version containing electrophoresis buffer and dye, which can be used directly after the reaction.

Product specifications:1 ml, 2ml, 5mL, etc., can be customized

Save condition:-30 ~ -15°C storage, ≤ 0°C transport

Scope of application:This product is suitable for PCR reactions using genomic DNA, cDNA, Plasmid DNA and DNA containing uracil as templates, and can also be used for colony PCR reactions.

2 × Tap Master Mix ForPAGE 说明书

Product number:PC 3117

Product Introduction:This product is expressed by Escherichia coli cloned with Thermus aquaticus DNA Polymerase gene and purified by multiple steps. It does not contain endonuclease, exonuclease and bacterial DNA. The product includes Taq DNA Polymerase, dNTPs, and an optimized buffer system that allows amplification with the addition of primers and templates, reducing pipetting operations and improving throughput and reproducibility of results. A version containing electrophoresis buffer and dye is available, and the accompanying 10 × Taq Buffer is specially optimized for polyacrylamide gel electrophoresis or agarose gel electrophoresis directly after the reaction.

Product specifications:1 ml

Save condition:-30 ~ -15°C storage, ≤ 0°C transport

Scope of application:This product is suitable for polyacrylamide gel electrophoresis of DNA polymerase, can be used for genomic DNA, cDNA, Plasmid DNA and uracil-containing DNA and colonies as the template of the PCR reaction.

2 × KeyPo Master Mix (Dye Plus)

Product number:PC 3089

Product Introduction:KeyPo DNA Polymerase is a new generation of high-fidelity DNA polymerase based on KOD DNA Polymerase. It maintains high amplification success rate and sensitivity, and can achieve rapid amplification of 5sec/kb. This product has a wide range of applicability to conventional templates, high GC systems (primers or templates) and uracil-containing templates. By adding two monoclonal antibodies that can inhibit its 5 '→ 3' polymerase activity and 3 '→ 5' exonuclease activity at normal temperature, this product can perform high-specific hot-start PCR reactions. At the same time, KeyPo Master Mix is a premixed 2 ×Mix, which can be amplified by adding templates and primers, which greatly simplifies the operation process.

Product specifications:1 ml, 2ml, 5mL, etc., can be customized

Save condition:-30 ~ -15°C storage, ≤ 0°C transport

Scope of application:This product is suitable for PCR reactions using genomic DNA, cDNA, Plasmid DNA and DNA containing uracil as templates.

Note:

1. The amplified length does not exceed 10kb;

2. In order to improve the success rate and yield of amplification, please use high-quality templates;

3. KeyPo DNA Polymerase has strong proofreading activity. Therefore, if TA cloning is required for the amplified product, DNA purification is recommended before adding A;

4. Primer design follows the principle of conventional PCR primer design, and the specific suggestions are as follows:

(1) The last base at the 3 'end of the primer is preferably G or C;

(2) The last 8 bases of the 3 'end of the primer should avoid continuous mismatch;

(3) Hairpin structure should be avoided at the 3 'end of the primer; Nanjing Sifan Biotechnology Co., Ltd. Artificial Intelligence Industrial Park, 266 Chuangyan Road, Nanjing Kirin Science and Technology Innovation Park

(4) The difference between the Tm values of the forward primer and the reverse primer is preferably not more than 1°C, and the Tm value is preferably adjusted to 55~65°C (Primer Premier 5 is recommended for the Tm value of the primer);

(5) The additional additional sequence of the primer, that is, the non-matching sequence with the template, should not be involved in the calculation of the Tm value of the primer;

(6) The GC content of primers was controlled between 40% and 60%;

(7) The overall distribution of primers A, G, C and T should be as uniform as possible, and areas with high GC or AT content should be avoided;

(8) Avoid complementary sequences of more than 5 bases within the primer or between the two primers, and avoid complementary sequences of more than 3 bases at the 3 'end of the two primers;

(9) After primer design, please use the NCBI BLAST function to search for primer specificity to avoid non-specific amplification.

Experimental process:

1. Reaction system

All operations should be carried out on ice, PCR-related components should be fully mixed after thawing, and put back to -20°C for storage after use.

Components Nuclease-free ddHzO 2xKeyPo master mix ( Dye Plus) Upstream primer (10 μM> Downstream primer (10 μM> Template

Volume Up to 50μL 25μL 2μL 2μL 1-5μL (not more than 1/10 of the total volume of PCR reaction system>

2. Reaction procedure

Temperature 98 ℃ TmNote 72 ℃

Time 10 Sec 5 Sec 5 Sec/kb

Number of cycles 25-38 cycles

Note: Please set the annealing temperature according to the Tm value of the primer. If the Tm value of the primer is ≥ 72°C, the annealing step can be deleted and the subsequent extension step (two-step PCR) can be directly performed. If necessary, it is recommended to find the optimum temperature for binding of the primer to the template by establishing a temperature gradient. In addition, the annealing temperature directly determines the amplification specificity, and if the amplification specificity is found to be poor, the annealing temperature can be appropriately increased.