T4 DNA Ligase 200μL - Uniscience

T4 DNA Ligase 200μL - Uniscience - Uniscience Corp.
Regular price $25.00 (UNI-R2081)

Overview

T4 DNA Ligase catalyzes the formation of phosphodiester bonds between juxtaposed 5' phosphate and 3' hydroxyl termini in double-stranded DNA using ATP as a coenzyme. Both blunt and cohesive end DNA ligation, as well as single-stranded nick repair of DNA, RNA and DNA/RNA, are possible via the T4 DNA ligase. This enzyme is supplied in a buffer of 10 mM Tris-HCl (pH 7.5), 50 mM KCl, 10 mM 2-mercaptoethanol, 0.1 mM EDTA
and 50% glycerol.

Biological Source

  • E. coli strain expressing a recombinant clone.

Molecular Weight

  • 68kDa.

Requirements

Mg , ATP and DTT. The optimum concentration of Mg is 10mM. Mn may be substituted for Mg but is only 25% as effective as Mg .

Inhibition

50% inhibition by greater than 150mM NaCl (activity measured at nicks). Other inhibitors include 0.2M K , Cs , Li , NH and 1mM spermine.

Inactivation

  • Heat to 70°C for 10 minutes.

Contents

  • T4 DNA Ligase (200 U/μl).
  • 10 X Ligation Buffer.

Applications

  • Cloning of restriction fragments.
  • Joining linkers and adapters to blunt-ended DNA.

Unit Definition

One unit of T4 DNA Ligase is defined as the amount of enzyme required to give 50% ligation of Hind III fragments of λ DNA (5´ DNA termini concentration of 0.12 μM, 300- μg/ml) in a total reaction volume of 20 μl in 30 minutes at 16°C in 1x T4 DNA Ligase Reaction Buffer.

Storage

  • Store at -20°C.

Storage Buffer

50 mM KCl, 10 mM Tris-HCl (pH 7.4), 0.1 mM EDTA, 1 mM DTT, 200 μg/ml BSA and 50% glycerol.

10X T4 Ligase Buffer

660mM Tris-HCl (PH7.6), 66mM MgCl , 100mM DTT, 1mM ATP.

Physical Purity

The purity is ≥90% as judged by SDS-polyacrylamide gel electrophoresis with Coomassie blue staining.

Standard Applications

We recommend using a 1:1, 1:3 or 3:1 molar ratio of vector:insert DNA when cloning a fragment into a plasmid vector. These ratios will vary with other types of vectors, for example, cDNA and genomic cloning vectors. The following example illustrates the conversion of molar ratios to mass ratios for a 3.0kb plasmid and a 0.5kb insert DNA
fragment.

Protocol

The following ligation reaction of a 3kb vector and a 0.5kb insert DNA uses a 1:1 vector:insert ratio. Typical ligation reactions use 100–200ng of vector DNA.

Download Datasheet


Overview

T4 DNA Ligase catalyzes the formation of phosphodiester bonds between juxtaposed 5' phosphate and 3' hydroxyl termini in double-stranded DNA using ATP as a coenzyme. Both blunt and cohesive end DNA ligation, as well as single-stranded nick repair of DNA, RNA and DNA/RNA, are possible via the T4 DNA ligase. This enzyme is supplied in a buffer of 10 mM Tris-HCl (pH 7.5), 50 mM KCl, 10 mM 2-mercaptoethanol, 0.1 mM EDTA
and 50% glycerol.

Biological Source

  • E. coli strain expressing a recombinant clone.

Molecular Weight

  • 68kDa.

Requirements

Mg , ATP and DTT. The optimum concentration of Mg is 10mM. Mn may be substituted for Mg but is only 25% as effective as Mg .

Inhibition

50% inhibition by greater than 150mM NaCl (activity measured at nicks). Other inhibitors include 0.2M K , Cs , Li , NH and 1mM spermine.

Inactivation

  • Heat to 70°C for 10 minutes.

Contents

  • T4 DNA Ligase (200 U/μl).
  • 10 X Ligation Buffer.

Applications

  • Cloning of restriction fragments.
  • Joining linkers and adapters to blunt-ended DNA.

Unit Definition

One unit of T4 DNA Ligase is defined as the amount of enzyme required to give 50% ligation of Hind III fragments of λ DNA (5´ DNA termini concentration of 0.12 μM, 300- μg/ml) in a total reaction volume of 20 μl in 30 minutes at 16°C in 1x T4 DNA Ligase Reaction Buffer.

Storage

  • Store at -20°C.

Storage Buffer

50 mM KCl, 10 mM Tris-HCl (pH 7.4), 0.1 mM EDTA, 1 mM DTT, 200 μg/ml BSA and 50% glycerol.

10X T4 Ligase Buffer

660mM Tris-HCl (PH7.6), 66mM MgCl , 100mM DTT, 1mM ATP.

Physical Purity

The purity is ≥90% as judged by SDS-polyacrylamide gel electrophoresis with Coomassie blue staining.

Standard Applications

We recommend using a 1:1, 1:3 or 3:1 molar ratio of vector:insert DNA when cloning a fragment into a plasmid vector. These ratios will vary with other types of vectors, for example, cDNA and genomic cloning vectors. The following example illustrates the conversion of molar ratios to mass ratios for a 3.0kb plasmid and a 0.5kb insert DNA
fragment.

Protocol

The following ligation reaction of a 3kb vector and a 0.5kb insert DNA uses a 1:1 vector:insert ratio. Typical ligation reactions use 100–200ng of vector DNA.

Download Datasheet

Warranty Information

All products we sell are covered solely by the manufacturer's warranty. Warranty information is included with the product and is subject to the conditions set forth by the manufacturer. We include such information in the product details for each item, when ever available. The manufacturer warrants their products are free from defects in material and workmanship. Repair or replacement may be issued for any product which is found to be defective under the terms of this warranty. Wholesale Point will assist with warranty issued within the first 14 days of product purchase. After that time the product manufacturer must be contacted directly to address all warranty issues. In many cases, the manufacturer can diagnose and resolve product concerns over the phone. For more information or to obtain contact information for the manufacturer, please call us +1(786) 420-2018.

Purchasing & Delivery

We take pride in making sure your order gets out quickly. We carefully check every item before shipping and we give extra care to our packaging to make sure every item arrives safely. Orders normally ship via UPS Ground, FedEx, or USPS within 1-2 days when in stock. Some items are drop shipped from the manufacturer and can take a little longer. For your convenience, whenever we ship an order, we electronically mail a shipping confirmation notice including the tracking number from UPS, FedEx, USPS, or a local delivery service.

Drop Ship Orders

Orders that are drop shipped from the manufacturer are subject to the manufacturer's availability and shipping schedule. Some items are special order or built-to-order and can take an average of 5-7 business days to ship. We make reasonable attempts to provide and display a shipping lead time on each product we sell. If you have any questions about our shipping policy or would like to know the availability or lead time on a specific drop ship product please feel free to contact us via our Contact Us Page. If you would like a drop shipped item expedited, please place your order by phone to guarantee your order can be expedited.

Payment Policy

We accept Visa, MasterCard, PayPal, American Express, and Discover.

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