商品编号


PAPY-40



品牌


MCLab



公司


MCLAB.Inc



公司分类


Reverse Transcriptase and RNA Polymerases



Concentration

600 U/ul


Size

240,000 units

商品信息

Poly(A) Polymerase from Yeast works more efficiently than
E. coli
Poly(A) Polymerase for RNA oligonucleotide-labeling and poly(A) tailing.




Description:
Poly(A) Polymerase catalyzes the template independent of the addition of AMP from ATP to the 3'-end of RNA. Poly(A) works more competently than
E. coli
poly(A) polymerase for RNA oligonucleotide-labeling and poly(A) tailing. Less incubation time is required for the yeast enzyme. This enzyme labels both long and short substrates. Poly(A) polymerase preferentially labels longer RNA-molecules whereas short RNA-molecules are labeled more efficiently by T4 RNA ligase. The reaction requires Mn
2+
or Mg
2+
, ATP as substrates, and any RNA containing 3'-hydroxyl termini as primers. Longer RNA molecules are somewhat better primers than short oligomers. Substitution of cordycepin 5'-triphosphate (3'-dATP) for ATP results in the addition of a single 3'-dA residue to the ends of the RNA, a useful technique for labeling RNA at the 3'-end.
Application:
- Labeling the 3'-ends of RNA with ATP or cordycepin
- Poly(A) tailing of RNA for cloning or affinity purification
- Preparing a priming site for
CDN
A synthesis using oligo-dT
- Enhancing translation of RNA transferred into eukaryotic cells
Source:
An
E. coli
strain that carries the cloned Poly(A) Polymerase gene from (
Saccharomyces cerevisiae
).
Specific Activity:
>20,000 U/mg
Unit Definition:
One unit is the amount of enzyme which incorporates 1 pmol AMP into acid-insoluble material at 37°C in 1 min.
5x Poly(A) Polymerase Reaction Buffer:
100 mM Tris-HCl, pH 7.0, 3.0 mM MnCl
2
, 0.1 mM EDTA, 1 mM DTT, 500 ?g/ml Acetylated BSA, 50% Glycerol.
Storage Buffer:
20 mM Tris-HCl (pH 8.0), 50 mM KCl, 0.5 mM DTT, 50% Glycerol.
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 ?l reaction. After incubation at 37°C for 10 min, acid insoluble r
ADI
oactivity 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.
Heat Inactivation:
65°C for 20 minutes
Recommended Storage Condition:
-20?C
References:
1. Sippel, A. E. (1973) Eur. J. Biochem. 37, 31-40.
2. Edmonds, M. (1982) in The Enzymes, 3rd edition, ed. P. D. Boyer (Academic Press, New York) 15, 217-244.
3. Gething, M. J., Bye, J., Skehel, J. and Waterfield, M. (1980) Nature 287, 301-306.
4. Sano, H. and Feix, G. (1976) Eur. J. Biochem. 71, 577-583.