Experimental design, in a 25μl system, using the same amount (0.1ng) of pCMV-Cre-EGFP as the template, using different amounts of this product or Bst DNA polymerase of Competitor, primer: 1.6µM FIP/BIP, 0.2µM F3/B3, 0.4µM Loop F/B, 1.4mM dNTP each, MgSO4 was added to 1X Bst Reaction Buffer (2mM MgSO4) until the final concentration reached 8mM, and incubated at 65ºC for 1 hour. Inactivation was heated at 80ºC for 20 minutes and then tested by 2.0% agarose gel electrophoresis.
As shown in the figure, this product has comparable enzyme activity compared with Competitor N's products.
Lane 1 UA070064- Bst DNA Polymerase, Full length 5U
Lane 2 Negative Control-1(negative control with no added enzyme only)
Lane 3 Negative Control-2(only negative controls without templates)
Lane 4 competing product N 5U
Bst DNA Polymerase, Full length
Bst DNA Polymerase, Full length
Product Details
Product Details
Product Specification
| Synonyms | DNA polymerase I |
| Expression System | E.coli |
| Molecular Weight | 100kDa (Reducing) |
| Purity | ＞95% by SDS-PAGE |
| Tag | His Tag |
| Storage Buffer | 50 mM KCl、10 mM Tris-HCl、0.1 mM EDTA、1 mM DTT、0.1% Triton® X-100、50% Glycerol、pH 7.1 @ 25°C |
| Stability & Storage | Store at -25 ~ -15℃ for 2 years |
| Reference | 1. Genetic Analysis: Biomolecular Engineering, 1996, 12(5-6):185-195. |
Background
Bst DNA Polymerase, Full Length is the full length polymerase from Bacillus stearothermophilus. It has 5´ → 3´ polymerase and double-strand specific 5´ → 3´ exonuclease activity, but lacks 3´ → 5´ exonuclease activity. It can be used to isothermal DNA amplification and Primer extension.
Components
50 mM KCl、10 mM Tris-HCl、0.1 mM EDTA、1 mM DTT、0.1% Triton® X-100、50% Glycerol、pH 7.1 @ 25°C
10* Reaction Buffer: 200 mM Tris-HCl、100 mM (NH4)2SO4、100 mM KCl、20mM MgSO4、0.1% Tween® 20、pH 8.8@25°C
Magnesium Sulfate (MgSO4) Solution：100mM MgSO4
Protocol
Incubate the following reaction at 65°C for 30–60 minutes
Component |
Final Concentration |
10X Isothermal Amplification Buffer II |
1X (contains 2 mM MgSO4) |
MgSO4 (100 mM) |
6 mM (8 mM total) |
dNTP Mix (10 mM) |
1.4 mM each |
FIP/BIP Primers (25X) |
1.6 µM |
F3/B3 Primers (25X) |
0.2 µM |
LoopF/B Primers (25X) |
0.4 µM |
Bst DNA Polymerase (8,000 U/ml) |
320 U/ml |
DNA or RNA Sample |
> 10 copies or more |
Nuclease-free Water |
to 25 µl |
Total Reaction Volume |
25 µl |
Guidelines
Bst DNA Polymerase does not exhibit 3´→ 5´ exonuclease activity.
Reaction temperatures above 70°C are not recommended.
Cannot be used for thermal cycle sequencing or PCR.
Unit Definition
Picture
Picture
Bioactivity
SDS-PAGE
2μg (R: reducing condition, N: non-reducing condition).
SEC-HPLC
