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E. coli Poly(A) Polymerase

Catalog # Concentration Size List Price Quantity Your Price
M0276L 5000 units/ml 500 units $467.00
$420.30
M0276S 5000 units/ml 100 units $116.00
$104.40
Catalog # Size List Price Your Price
M0276L 500 units $467.00
$420.30
M0276S 100 units $116.00
$104.40
Catalog #
Qty:
 
*On-line ordering is for Canadian customers only. Web pricing is applicable only to orders placed online at www.neb.ca

Poly(A) Polymerase catalyzes the template independent addition of AMP from ATP to the 3´ end of RNA

  • Isolated from a recombinant source
  • Applications include 3' labeling of RNA with ATP or Cordycepin and poly(A) tailing of RNA for cloning or affinity purification
  • Enhances translation of RNA transferred into eukaryotic cells
  • Tested for the absence of DNases and RNases
Featured Videos
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  • Monarch_MiniprepProtocol_VideoThumb
    Avoiding RNase Contamination
Poly(A) Polymerase catalyzes the template independent addition of AMP from ATP to the 3´ end of RNA.

Figure 1. Analysis of capped and polyadenylated RNA
Figure 1
A. Agilent Bioanalyzer® analysis of capped and polyadenylated RNA. Longer tails are produced by increasing the enzyme concentration in the reaction. Calculated A-tail lengths are indicated over each lane. Lanes: L: size marker,1: No poly-A tail, 2: 5 units, 3 :15 units, 4 : 25 units of E. coli Poly(A) Polymerase. B. Effect of enzymatic A-tailing on the luciferase reporter activity of CLuc mRNA
Product Source
An E. coli strain that carries the cloned Poly(A) Polymerase gene from E. coli (1).
Reagents Supplied

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration
  E. coli Poly(A) Polymerase M0276SVIAL -20 1 x 0.02 ml 5,000 units/ml
  Poly(A) Polymerase Reaction Buffer B0276SVIAL -20 1 x 1.5 ml 10 X
  Adenosine-5'-Triphosphate (ATP) B0756AVIAL -20 1 x 0.2 ml 10 mM
  E. coli Poly(A) Polymerase M0276LVIAL -20 1 x 0.1 ml 5,000 units/ml
  Poly(A) Polymerase Reaction Buffer B0276SVIAL -20 1 x 1.5 ml 10 X
  Adenosine-5'-Triphosphate (ATP) B0756AVIAL -20 1 x 0.2 ml 10 mM
Application Features
  • Labeling of RNA with ATP or cordycepin
  • Poly(A) tailing of RNA for cloning or affinity purification
  • Enhances translation of RNA transferred into eukaryotic cells.

Properties & Usage

Unit Definition

One unit is defined as the amount of enzyme that will incorporate 1 nmol of AMP into RNA in a 20 µl volume in 10 minutes at 37°C.

Reaction Conditions

1X Poly(A) Polymerase Reaction Buffer
Supplement with 1 mM ATP
Incubate at 37°C

1X Poly(A) Polymerase Reaction Buffer
50 mM Tris-HCl
250 mM NaCl
10 mM MgCl2
(pH 8.1 @ 25°C)

Storage Buffer

20 mM Tris-HCl
300 mM NaCl
1 mM DTT
1 mM EDTA
50% Glycerol
0.1% (w/v) Triton® X-100
pH 7.5 @ 25°C

Unit Assay Conditions

1X Poly(A) Polymerase Reaction Buffer, 1 mM rATP and 500 ng 5´ FAM labeled poly A 20-mer RNA in a 20 µl reaction. After incubation at 37°C for 10 minutes the length of the poly(A) addition is determined either by gel electrophoresis or with an automated capillary DNA sequencer. In this assay 5 units of enzyme add approximatley 60 to 80 adenosines to the RNA primer. In these conditons 20 units of enzyme will deplete the rATP.

Notes
  • rATP is not included in the buffer supplied.
References
  • Cao, G.J. and Sarkar, N. (1992). Proc.Natl. Acad. Sci. USA. 89, 10380-10384.
FAQs
Tech Tips

  • E. coli Poly (A) Polymerase (M0276)

    Vortex tube gently before use.

    Poly(A) Polymerase can be inactivated by adding EDTA at a final concentration of 10 mM to the reaction.

    Poly (A) Polymerase is compatible with the M-MuLV reverse transcriptase buffer. If the intention is to carry out the polyadenylation step followed by reverse transcription, we recommend to set-up these reactions in two sequential steps. The polyadenylation reaction can be done first using the reverse transcriptase buffer. This can be followed by addition of the components necessary for reverse transcription.

    We strongly recommend wearing gloves, using nuclease-free tubes and reagents, and thoroughly cleaning pipettes and bench surfaces to avoid RNase contamination.

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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