Quick Blunting™ Kit

Application Features

Prepare sheared, nebulized or restriction enzyme digested DNA for blunt-ended ligation into a plasmid, cosmid, fosmid or BAC vector

Prepare PCR products for efficient blunt-end cloning

Companion Products

• Quick Blunting™ and Quick Ligation™ Kits
• Quick T4 DNA Ligase
• Monarch® Plasmid Miniprep Kit 
• Monarch® DNA Gel Extraction Kit
• Monarch® PCR & DNA Cleanup Kit (5 μg)

1. Restriction enzyme digested DNA can be blunted directly without purification. The Blunt Enzyme Mix has been optimized in Blunting Buffer, but is also active in NEBuffers 1,2,3 and 4, as well as BamHI, EcoRI and DpnII unique buffers when supplemented with dNTPs and dithiothreitol. There is a small reduction in ligation fidelity in these buffers. Transformation efficiency is lowest in NEBuffer 1 where the total yield is about 50% of optimum.
2. ATP is not necessary for T4 Polynucleotide Kinase activity in the kit. The dATP and dTTP in the dNTP mix act as phosphate donors.
3. The blunted reaction must be purified prior to phosphatase treatment by using a commercial purification kit, phenol extraction/ethanol precipitation, or gel electrophoresis.
4. PCR generated DNA must be purified before blunting by using a commercial purification kit, such as Monarch® PCR & DNA Cleanup Kit (NEB #T1030), phenol extraction/ethanol precipitation, or gel electrophoresis.

1. Blunting protocol for NEB PCR Cloning Kit
2. Protocol for the Quick Blunting Kit (E1201)
3. Transformation Protocol
4. Quick Ligation Protocol (M2200)

• Blunting Selection Chart

• NEBcloner®

1. Which ligase is recommended to ligate DNA treated with the Quick Blunting Kit?
2. Can I use regular ligase to ligate my blunted product?
3. I've blunted my DNA and now need to dephosphorylate the reaction with Antarctic Phosphatase. Do I need to purify the reaction?
4. Does the PCR product need to be purified before blunting with the Quick Blunting Kit? What methods can be used to purify the product?
5. I've digested my DNA and now want to blunt directly without purifying, can I add the blunting reagents directly to the restriction digest?
6. I've blunted my sonicated gDNA for 15 minutes instead of the recommended 30 minute incubation time, will the reaction still work?
7. I've accidentally skipped the heat-kill step after the blunting reaction. Will the ligation still work?

• EpiMark® Methylated DNA Enrichment Kit Troubleshooting Guide

Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.

Specifications

The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]

• E1201S_L_v1

The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]

• E1201S_L_v1_0101801
• E1201L_v1_10013484
• E1201L_v1_10013485
• E1201L_v1_10038381
• E1201S_v1_10038380
• E1201S_v1_10053797
• E1201S_v1_10058037
• E1201S_v1_10062856
• E1201L_v1_10053799
• E1201L_v1_10062855
• E1201S_v1_10080108
• E1201L_v1_10085502
• E1201L_v1_10102455
• E1201S_v1_10107427

The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

• Blunting Enzyme Mix
• 10X Blunting Buffer
• Deoxynucleotide Solution Mix (1 mM)

This product is covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB).

While NEB develops and validates its products for various applications, the use of this product may require the buyer to obtain additional third party intellectual property rights for certain applications.

For more information about commercial rights, please email us at busdev@neb.com.

This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.