Tth DNA Polymerase

Cat. No.: T292976

GRADE & PURITY Recombinant, Suitable for molecular biology, BioReagent, PCR Reagent, EnzymoPure™, for DNA and RNA applications, From Thermus thermophilus

Overview

Tth DNA polymerase is a thermostable enzyme with a molecular weight of 94 kDa. It is recombinantly expressed in Escherichia coli from the gene of Thermus thermophilus HB‑8. The enzyme performs DNA replication at 74 °C and has a half‑life of 20 min at 95 °C. In the presence of magnesium ions, Tth DNA polymerase catalyzes the polymerization of nucleotides in the 5′→3′ direction to form double‑stranded DNA. In the presence of manganese ions, it can also carry out nucleotide polymerization in the 5′→3′ direction using RNA as a template. This enzyme also possesses 5′→3′ exonuclease activity. Tth DNA polymerase can be applied to PCR, RT‑PCR, reverse transcription, and primer‑extension reactions at elevated temperatures.

Contents & Storage:

Component No.	Component Name	Appearance	50μL	250μL	1ml	Storage
T292976A	Tth DNA Polymerase	Liquid	50μL	250μL	1ml	-20℃.
T292976B	2×Tth One Step RT-PCR Buffer	Liquid	1.25ml	6.25ml	25ml	-20℃.
T292976C	Mn²⁺ (50×)	Liquid	50μL	250μL	1ml	-20℃.

Note: 2×Tth One Step RT‑PCR Buffer contains Mg²⁺.

Scope of application:

PCR amplification, RT‑qPCR amplification.

Unit definition:

One unit is defined as the amount of enzyme required to catalyze the incorporation of 10 nmol dNTPs into TCA‑insoluble material within 30 min at 70 °C.

Precautions:

1. Dissolve and mix all reaction‑required solutions at room temperature or 4 °C, then place them on an ice bath or in an ice box.

2. It is recommended to aliquot the reaction mixture to avoid repeated freeze‑thaw cycles.

3. Excessive template DNA is liable to produce non‑specific PCR products.

Procedure:

Reaction System Setup:

Component	Volume Added	Final Concentration
2×Tth One Step RT-PCR Buffer	25μl	1×
RNasin (40U/μl)	1μl	/
Forward Primer (10μM)	1μl	0.5μM
Reverse Primer (10μM)	1μl	0.5μM
Probe (10μM)	0.25μl	0.5μM
Mn²⁺ (50×)	1μl	1×
Tth DNA Polymerase	1μl	0.1U/μ1
Template DNA	5μl	/
ddH₂O	To 50μl	/
Final Volume	50μl	/

Reaction procedure:

Step Name	Temperature	Time	Cycles
Heat Activation	95℃	20min 30s	1
Annealing	60℃	30s	1
Heat Activation	95℃	20min 30s	1
Denaturation	94℃	15s	40
Annealing	55℃	40s
Hold	4℃	∞	1

Specifications

Unit definition

At 70°C, the amount of enzyme required to catalyze the incorporation of 10nmol dNTPs into TCA-insoluble substances within 30 minutes is one unit.

Storage and Shipping

Storage

Store at -20°C,Avoid repeated freezing and thawing

Shipped In

Ice chest + Ice pads

Stability And Storage

Store at -20℃ long term (24 months). Avoid repeated freeze-thaw cycles.

Contents & Storage

Component No.	Component Name	Appearance	50μL	250μL	1ml	Storage
T292976A	Tth DNA Polymerase	Liquid	50μL	250μL	1ml	-20℃.
T292976B	2×Tth One Step RT-PCR Buffer	Liquid	1.25ml	6.25ml	25ml	-20℃.
T292976C	Mn²⁺ (50×)	Liquid	50μL	250μL	1ml	-20℃.

Note: 2×Tth One Step RT‑PCR Buffer contains Mg²⁺.