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Enzymes for Molecular Biology

Enzymes for Molecular Biology: Enzymes for Molecular Biology

RNA Poly A Polymerase

Poly A RNA polymerase adds poly A tails to the 3? hydroxyl terminus of RNA and is useful for cloning purposes. 3?-end labelling RNA with a[32P]-cordycepin triphosphate and determining molar concentration of polyadenylated RNA. All preparations are assayed for contaminating ribonuclease activity

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Description

Source

Escherichia coli

Applications

  • Adds poly A tails to the 3’-hydroxyl terminus of RNA (1)
  • Labels 3’-ends of RNA with a [α-32P] cordycepin triphosphate (2)
  • Ideal for determining molar concentration of polyadenylated RNA (3)

Unit Definition

One unit is the amount of enzyme required to incorporate 1.0 nmol of labeled ATP into acid insoluble form for 10 minutes at 37°C under the stated assay conditions

Concentration

1 Unit/µl

Storage Buffer

25 mM Tris-HCl (pH 7.9) 
0.1 mM DTT
0.01% (v/v) Triton X-100™
1.0 mM EDTA
0.5 M NaCl50% (v/v) glycerol

Assay Conditions

50 mM Tris-HCl (pH 7.9)
10 mM MgCl2
2.5 mM MnCl2 
250 mM NaCl
25 µg tRNA
0.1 mM [α-32P] ATP
50 μg bovine serum albumin
Reaction volume of 100 µl

Storage Conditions

Store at -20°C

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Protocols

Download here

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References

(1) Sippel, A.E. (1973) Eur. J. Biochem. 37, 31-40 (2) Beltz, W.R. and Ashton, S.H. (1982) Fed. Proc. 41, 1450, Abstract 68996 (3) Krug, M.S., and Berger, S.L. (1986) Anal. Biochem. 153, 315-323

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