T5 exonuclease degrades DNA along with the 5'→3' direction, which degrades double-stranded DNA, single-stranded DNA, and nicked plasmid DNA. It can degrade DNA both from the 5'-end and from the cut or notch of linear or circular double-stranded DNA, but it cannot degrade superhelical double-stranded DNA. Based on the above characteristics, T5 exonuclease can be applied to the Gibson assembly.
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Cat. No.: T5EX-1k (for 1,000U)
Cat. No.: T5EX-5k (for 5KU)
Cat. No.: T5EX-20k (for 20KU)
T5 Exonuclease is a type of exonuclease that degrades double-stranded or single-stranded DNA in the 5'→3' direction. T5 Exonuclease can initiate digestion from the 5' end of single-stranded or double-stranded DNA and can also start digestion from the gaps or nicks in linear or circular double-stranded DNA.
T5 Exonuclease cannot degrade supercoiled double-stranded DNA, and its activity in degrading single-stranded DNA can be inhibited by reducing the Mg2+ concentration in the reaction buffer to below 1mM. Based on these characteristics, T5 Exonuclease is commonly used in Gibson Assembly.
Gibson Assembly is a technique used to efficiently connect multiple overlapping DNA fragments under isothermal conditions. The basic principle of Gibson Assembly can be summarized in three steps: (1) T5 DNA exonuclease initiates digestion from the 5' end of DNA fragments, generating complementary single-stranded 3' overhangs that facilitate annealing of complementary ends; (2) DNA polymerase fills in the gaps created by annealing; (3) DNA ligase connects the assembled fragments at the nicks, resulting in a complete double-stranded DNA.
T5 Exonuclease is commonly used in Gibson Assembly, degradation of linear single-stranded, double-stranded, or nicked plasmid DNA, removal of incomplete ligation products from circular double-stranded DNA, degradation of linear and nicked plasmid DNA to obtain high-purity supercoiled plasmid DNA, removal of denatured plasmid DNA generated during alkaline extraction of plasmids, and improvement of transfection efficiency of small-scale purified plasmid cDNA libraries.
Purified from E. coli strains expressing the T5 bacteriophage D15 gene plasmid.
One unit is defined as the amount of enzyme required to produce 1 nmol of acid-soluble deoxyribonucleotide from double-stranded DNA in 30 minutes at 37°C in a total reaction volume of 50 µl.
The enzyme does not contain any DNA endonucleases or exonucleases other than T5 Exonuclease, RNAse, or phosphatases.
T5 Exonuclease can be inactivated by adding EDTA to a final concentration of at least 11mM or by using DNA loading buffer containing SDS (final SDS concentration is 0.08%).
The minimum shelf life is 3 years at -20°C.
T5 Exonuclease is a selectively acting DNA exonuclease that exhibits different reaction activities with different DNA substrates. Therefore, when digesting specific substrates, it is important to properly control the enzyme amount and reaction time.
The optimal reaction temperature for T5 Exonuclease is 37°C, but it also has some activity at 50°C, making it suitable for use in Gibson Assembly.
T5 Exonuclease also exhibits activity in regular PCR buffers.
This product is intended for scientific research by professionals only and must not be used for clinical diagnosis or treatment, food or drugs, and should not be stored in regular residential areas.
For your safety and health, please wear lab coats and disposable gloves during handling.
Only for research and not intended for treatment of humans or animals