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Products >  PCR_Products >  High_Fidelity_PCR >  PrimeSTAR_HS_DNA_Polymerase

PrimeSTAR HS DNA Polymerase

PrimeSTAR HS DNA Polymerase, the original enzyme in the PrimeSTAR series, is a novel PCR enzyme that provides high fidelity and accuracy during amplification of large products (8.5 kb for human genomic DNA; 22 kb for lambda DNA). This hot start PCR enzyme can be useful during highly demanding cloning (e.g., amplification of cDNA libraries) and sequencing applications. The fidelity of PrimeSTAR HS DNA Polymerase has been calculated by direct sequencing analysis. Such analysis provides a more realistic estimate of the actual frequency of incorporation errors compared with other commonly-used indirect phenotypic analyses such as lacl. Calculation methods such as lac1, which are often used by other enzyme suppliers, are prone to error due to inherent lethal and silent mutations.

PrimeSTAR HS DNA Polymerase also offers excellent amplification efficiency and a short reaction time. Because PrimeSTAR HS is a hot start PCR enzyme, false initiation events during reaction assembly are minimized, reducing experimental background.

PrimeSTAR HS DNA Polymerase possesses extremely high priming efficiency. Therefore, highly specific amplifications can be achieved using short annealing times of just 5-15 seconds.

PrimeSTAR HS (Premix): The PrimeSTAR HS (Premix) is an optimized mixture composed of 2X PrimeSTAR HS DNA Polymerase, reaction buffer and dNTP mixture. This is useful for high-throughput applications because the 2X premixed reaction format reduces contamination risk.

Note regarding fidelity comparisons: Takara uses sequencing analysis to determine incorporation error rate because most researchers who require high fidelity perform the following reactions: PCR, cloning and sequencing. Determination of error rate by direct sequence analysis, therefore, corresponds closely to the method that most researchers would use during their own applications. Please note that values obtained using different fidelity measurement methods (e.g., direct sequencing, Kunkel method, Cline method, lacI method) do not yield directly comparable data.

At-A-Glance Documents Images & Data Resources

Features

  • High accuracy and strong exonuclease activity, resulting in an extremely low error rate of only 12 errors per 250 kb.
  • Higher amplification efficiency compared with standard Taq polymerase.
  • Excellent performance even with GC-rich templates.
  • High tolerance to varying reaction conditions (single PCR cycling protocol can be used to amplify products of varying sizes).
  • Amplification of targets up to 8.5 kb with human genomic DNA, 10 kb with E. coli genomic DNA, and 22 kb with lambda DNA.
  • Fast reaction time due to increased priming efficiency.
  • Hot start PCR enzyme prevents false initiation events during reaction assembly due to mispriming or primer digestion.

Applications

  • High fidelity PCR
  • Amplification of cDNA libraries
  • cDNA cloning for protein expression
  • Site-directed mutagenesis and mutant genotyping (e.g., SNP analysis)

Components

R010A

PrimeSTAR HS DNA polymerase100 µL
5X PrimeSTAR Buffer (Mg2+)2 x 1 mL
dNTP Mixture800 µL
 

R040A Premix Version

PrimeSTAR HS Premix*5 X 500 µL

*Contains PrimeSTAR HS DNA Polymerase (1.25 units/25 µL); 2X dNTP Mixture (0.4 mM); 2X PrimeSTAR Buffer (2mM Mg2+).

Purity

Nicking, endonuclease and exonuclease activity are not detected following incubation of 0.6 µg of supercoiled or lambda-Hind III digested pBR322 DNA with 10 units of enzyme for 1 hour at 74°C.

Storage

–20°C. Premix: Shipped and stored at –20°C. Avoid repeated freeze-thaw and vigorous stirring. Once thawed, dispense into PCR tubes and store at –20°C.

Performance

Enzymatic performance was confirmed by single fragment PCR using both lambda DNA (amplified fragments: 8, 10, 12, 15 kb) and human genomic DNA (amplified fragment: 0.5, 1, 2, 4, 6, 8 kb) as templates.

PCR Products

A significant percentage of PCR products obtained using PrimeSTAR HS DNA polymerase will have blunt ends. As a result, PCR products can be directly cloned into blunt-ended vectors. If necessary, phosphorylate PCR products before cloning.

References

1. Dare, A.P. et al. (2008) Plant Methods 4:17.

2. Belton, Jr., R.J. et al. (2008) J. Biol. Chem. 283:17805-17814.

3. Nakatsubo, T. et al. (2008) J. Biol. Chem. 283:15550-15557.

4. Bolliger, M.F. et al. (2008) Proc. Natl. Acad. Sci. USA 105:6421-6426.

5. Seema, C. et al. (2008) J. Bacteriol. 190:2987-2996.

6. Russell, R.S. et al. (2008) Proc. Natl. Acad. Sci. USA 105:4370-4375.

7. Inoue, J. et al. (2007) Eukaryot. Cell 6:1925-1932.

8. Kawasaki, T. et al. (2007) J. Bacteriol. 189:5792-5802.

9. Daquinag, A. et al. (2007) Mol. Cell. Biol. 27:633-650.

10. Okahara, F. et al. (2006) Mol. Biol. Cell. 17:4888-4895.

11. Kasai, K. et al. (2006) J. Bacteriol. 188:7111-7122.




 
 
Products
Cat. # Product Package Size Price License # of Units Select
R010A PrimeSTAR® HS DNA Polymerase 250 Units $185.00 License Statements
R010B PrimeSTAR® HS DNA Polymerase 1,000 Units $618.00 License Statements
R040A PrimeSTAR® HS DNA Polymerase (premix) 100 Rxns $138.00 License Statements
 

 


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