The A-Plus™ Poly(A) Polymerase Tailing Kit uses ATP as a substrate for template-independent addition
of adenosine monophosphates to the 3'-hydroxyl termini of RNA. The standard protocol produces a
poly(A)-tail length of ~150 b on 60 μg of RNA. Polyadenylation increases the stability of RNA in
eukaryotic cells and enhances its ability to be translated after transfection or microinjection.1-3 A Poly(A)
tail is useful to provide a priming site for first-strand cDNA synthesis in certain applications, and can be
used to end-label4 or quantify5 mRNA.
The nuts and bolts of translational efficiency and the "closed-loop" model
Store at –20°C in a freezer without a defrost cycle. Do not store at –70°C.
|A-Plus™ Poly(A) Polymerase Tailing Kit Contents (50 reactions)
|A-Plus™ Poly(A) Polymerase, 4 U/μl
in 50% glycerol, 50 mM Tris-HCl, pH 7.5, 0.5 M NaCl, 1 mM
dithiothreitol (DTT) and 0.1 mM EDTA.
|10X A-Plus™ Poly(A) Tailing Buffer
0.5 M Tris-HCl, pH 8.0, 2.5 M NaCl and 100 mM MgCl2.
|10 mM ATP
||2 x 1.4 ml
Materials Required, but not Supplied
• Materials or kits for purification of the RNA product. (For suggestions, see RNA Purification, page 4)
• RNase-free TE Buffer (10 mM Tris-HCl, pH 7.5, 1 mM EDTA)
• Optional: TE saturated phenol/chloroform, 0.5-1 M EDTA, ScriptGuard™ RNase Inhibitor
One unit of A-Plus Poly(A) Polymerase converts 1 nmole of ATP into acid-insoluble material in 10
minutes at 37oC under standard assay reaction conditions.
The A-Plus Poly(A) Polymerase Tailing Kit is functionally tested in 1X A-Plus Poly(A) Tailing Buffer
with 1 mM ATP, a 1.4 kb RNA transcript and varying amounts of A-Plus Poly(A) Polymerase.
Contaminating Activity Assays
All components of the A-Plus Poly(A) Polymerase Tailing Kit are free of detectable RNase and DNase
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