Taq DNA polymerase (+dNTPs), with Enhancer for high GC template (200 units)
Taq DNA polymerase (Enhancer for high GC template)
Description
Thermus aquaticus DNA polymerase (Taq DNA polymerase) was expressed in E. coli in large quantities and highly purified. The enzyme has thermostable DNA polymerase activity and the MW is 94 kDa. This enzyme kit is especially suitable for PCR reactions with high GC template due to Ehancer for high GC templates and Robust buffer.
Application
- High-throughput PCR
- Colony PCR
- Incorporation of dUTP, dITP, and fluorescence-labeled nucleotides
- Primer extension
- Addition of a single nucleotide (adenosine) at the 3’-blunt ends (for cloning into TA vector)
Products
| Product ID | dNTP | Size | Link |
|
FNK-02-013
|
- | 200 units |  |
|
FNK-02-013-5
|
- | 5x200 units | |
|
FNK-02-003
|
+ | 200 units | |
|
FNK-02-003-5
|
+ | 5x200 units | |
General composition of PCR reaction mixture (total 50ul)
| Taq DNA polymerase (5 units/ul) | *0.25 ul |
| 10 x Standard Buffer (Taq) | 5 ul |
| 2.5mM (each) dNTPs | 4ul |
| Template | <500ng |
| Primer 1 | 0.2～1.0uM (final conc.) |
| Primer 2 | 0.2～1.0uM (final conc.) |
| Sterile distilled water | up to 50ul |
*Use of excess amount of enzyme is not recommended.
Storage Conditions
20mM Tris-HCl (pH 8.0), 100mM KCl, 0.1mM EDTA, 1mM DTT, 50% glycerol, 0.5% Tween20, 0.5% Igepal CA-630, Store at -20℃
Concentration
5 units/ul, where one unit is defined as the amount of enzyme that can incorporate 10 nmols of total dNTPs into an acid-insoluble material in 30 minutes at 74℃ when activated salmon sperm DNA was used as template/primer.
Quality AssurancePCR Test: Good amplification result was obtained in PCR reaction using λDNA as a template up to 14 kB (Fig.2).
Reagents Supplied with Enzyme
- 10 x Robust Buffer (Taq)
- 5 x GC Enhancer
Cautions for usage of Robust Buffer (Taq) without GC Enhancer
Robust Buffer induces maximum enzymatic activity. Therefore, cares should be taken to avoid production of undesirable smear bands in gel electrophoresis analysis by longer than optimal reaction time. We recommend about 5 to 10 seconds / kb elongation time for template up to 8 kb, and about 15 seconds / kb for up to 14 kb. We will recommend roughly the same elongation time to be set with 2-step PCR (shuttle PCR) and 3-step PCR. Extend the elongation time by short steps when amplification is not seen. The results of your experiments can be observed more rapidly by adopting 2-step PCR.
Protocols for PCR
Examples of PCR coditions without GC Enhancer for the amplification of various sizes of λDNA (Results shown in Fig.2)
Examples of PCR coditions with GC Enhancer for the amplification of the adenylate cyclaseA gene from Bordetella pertussis（ToHAMA I) genomic DNA (GCcontent 67%) (Results shown in Fig.3)
Fig.3 Effect of the Enhancer on the efficiecy of POR with high GC template (the adenylate cyclase
gene from Bordetella pertussis; 67% GC, 6 kb)
