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5´ DNA Adenylation Kit

Catalog # Concentration Size List Price Quantity Your Price
E2610L 50 µM 50 reactions $784.00
$705.60
E2610S 50 µM 10 reactions $196.00
$176.40
Catalog # Size List Price Your Price
E2610L 50 reactions $784.00
$705.60
E2610S 10 reactions $196.00
$176.40
Catalog #
Qty:
 
*On-line ordering is for Canadian customers only. Web pricing is applicable only to orders placed online at www.neb.ca
  • This kit contains Mth RNA Ligase (NEB #M2611A)
  • Ideal for generating high yields of 5´ adenylated DNA
  • Easy to use protocol with >95% efficiency
  • One step reaction with no need for gel purification
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    Avoiding RNase Contamination
The 5´ DNA adenylation Kit is a simple and efficient enzymatic method for generating 5´-adenylated DNA. The kit is optimized to produce the adenylated DNA with or without 3´-terminator. The 5´ DNA adenylation kit routinely generates greater than 95% conversion of pDNA to AppDNA(1). This highly efficient process eliminates the need for gel isolation of the product and increases overall yield.
Reagents Supplied

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration
  Mth RNA Ligase M2611AVIAL -20 1 x 0.02 ml 50 µM
  5' DNA Adenylation Reaction Buffer B2610SVIAL -20 1 x 1 ml 10 X
  Adenosine 5' Triphosphate N0757AVIAL -20 1 x 0.1 ml 1 mM
  Mth RNA Ligase M2611AAVIAL -20 1 x 0.1 ml 50 µM
  5' DNA Adenylation Reaction Buffer B2610SVIAL -20 1 x 1 ml 10 X
  Adenosine 5' Triphosphate N0757AVIAL -20 1 x 0.1 ml 1 mM
Features
  • One step reaction gives quantitative adenylation
  • Simpler than existing chemical and enzymatic methods
  • Reduces need for extensive purification of reaction product
  • 65°C reaction temperature reduces secondary structural concerns
  • Easily scalable from pmol to μmol range
Application Features
Enzymatic 5´-adenylation of single-stranded DNA linkers for next generation sequencing.

Properties & Usage



Notes
  • The adenylation reaction can be scaled up to 6X without a loss of efficiency, to a final concentration of 30 pmol of oligonucleotide and 30 pmol of Mth RNA Ligase per µl. The oligonucleotide can be purified by phenol extraction and alcohol precipitation or column chromatography to remove protein and ATP.
  • For substrates with unprotected 3´ termini increase concentration of ATP to 0.5 mM to prevent circularization and concatemerization.
  • The low turnover of the enzyme requires an approximately equimolar concentration of the enzyme and the oligonucleotide substrate.
  • Adenylated DNA linkers can be used for 3´-end ligation of RNA in cDNA library preparation for Next Generation sequencing protocols [3,4].
References
  • Zhelkovsky, A.M. and McReynolds, L.A. (2011). Nucl. Acids Res. 39(17): e117.
  • Torchia, C., Takagi, Y. and Ho, C.K. (2008). Nucleic Acids Res. 36, 6218-6227.
  • Hafner, M. et al. (2008). Methods. 44, 3-12.
  • Vigneault, F., Sismour, A.M. and Church, G.M. (2008). Nature Methods. 5, 777-779.
Quality Control Assay
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]
Certificate of Analysis
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]
Safety Data Sheets
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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.

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