Tth DNA Polymerase is a heat-resistant DNA polymerase discovered from the thermophilic bacterium Thermus thermophilus HB8. In the presence of Mg²⁺, this enzyme exhibits 5′-3′ DNA polymerase activity and 5′-3′ exonuclease activity, but lacks 3′-5′ exonuclease activity, making it widely applicable for PCR amplification reactions. In the presence of Mn²⁺, the enzyme demonstrates strong reverse transcriptase activity at temperatures of 55-70°C, making it suitable for one-step RT-PCR reactions. Specification Unit Definition: One unit (U) of activity is defined as the amount of enzyme that incorporates 10 nmol of total nucleotides into an acid-insoluble product in 30 minutes at 74°C, using activated salmon sperm DNA as the template/primer. Component No. Name Size 14607ES72 (250 U) 14607ES80 (1250 U) 14607-A Tth DNA Polymerase (5 U/μL) 50 μL 250 μL 14607-B 5×Tth RT-PCR Buffer 1 mL 4×1.25 mL 14607-C 10×Tth PCR Buffer (with Mg2+) 500 μL 2×1.25 mL 14607-D 50mM Mg(OAc)2 250 μL 1.25 mL 14607-E dNTP Mixture (10 mM each) 150 μL 750 μL Shipping and Storage Shippingwith dry ice. Store at -20°C, with a shelf life of 2 years. Precautions 1) If a small amount of precipitate appears in the buffer upon thawing, this is a normal phenomenon. Please invert and mix thoroughly before use. 2) For your safety and health, wear a lab coat and disposable gloves during operation. 3) This product is for research use only! Instructions Recommended PCR Reaction Procedure: 1. Preparation of the Reaction System Name Volume (μL) 10×Tth PCR Buffer (with Mg2+) 5 Tth DNA Polymerase (5 U/μL) 0.5 dNTP Mixture (10 mM each) 1 Upstream Primer 10μM 2 Downstream Primer 10μM 2 Template DNA 50 pg- 1μg ddH2O to 50 [Note]: The amounts of Tth DNA Polymerase, upstream and downstream primers, and dNTP Mixture can be adjusted according to specific experimental needs. 2. Cycle Setup Steps Temp(ºC) Time Cycles Pre-denaturation 94 30 sec-5 min 1 Denaturation 94 30 sec 35 Annealing 50-70 30 sec Extension 72 60 sec/kb Final Extension 72 10 min 1 [Note]: Reaction conditions can be adjusted according to specific experimental requirements. Recommended RT-PCR Reaction Procedure: 1. Preparation of the Reaction System Name Volume (μL) 5×Tth RT-PCR Buffer 10 Tth DNA Polymerase (5 U/μL) 1.5 50mM Mg(OAc)2 2.5 dNTP Mixture (10 mM each) 1.5 Upstream Primer 10μM 2 Downstream Primer 10μM 2 Template RNA 50 pg- 1μg ddH2O to 50 [Note]: The amounts of Tth DNA Polymerase, upstream and downstream primers, and dNTP Mixture can be adjusted according to specific experimental needs. 2. Cycle Setup Steps Temp (ºC) Time Cycles Reverse transcription 55-70 30 min 1 Pre-denaturation 94 30 sec-5 min 1 Denaturation 94 30 sec 35 Annealing 50-70 30 sec Extension 72 60 sec/kb Final Extension 72 10 min 1 [Note]: Reaction conditions can be adjusted according to specific experimental requirements.
