18018-010 SP6 RNA Polymerase
Y90109 5X SP6 Buffer
Y01500 10 mM DTT
One unit incorporates 1 nmol of labeled nucleotide into acid-precipitable material in 1 hour at 37ºC.
Storage Buffer: 5X SP6 Buffer:
50 mM Tris-HCl (pH 7.9) 0.2 M Tris-HCl (pH 7.9)
0.1 M NaCl 30 mM MgCl 2
0.1 mM EDTA 10 mM spermidine-(HCl) 3
14 mM 2-mercaptoethanol Refer to Functional Assay
50% (v/v) glycerol Conditions on reverse side for
0.1% (w/v) Triton® X-100 further details.
Store buffer at -20ºC.
Store 10 mM DTT at -20ºC.
This product has passed the following quality control assays: functional absence of exonuclease, endo-ribonuclease and DNA nicking activities; performance in a transcription reaction. The enclosed buffers were assayed with the enzyme and met quality control specifications.
Functional Assay Conditions:
2 μl 5X SP6 Buffer
2.5 μM [α- 32P]UTP (10 μCi at 400 Ci/mmol)
10 μM UTP
0.4 mM each ATP, CTP, GTP
1 mM DTT
0.2 μg linearized template DNA
15 units SP6 RNA Polymerase
Reaction Volume: 10 μl
Incubation: 60 minutes at 37°C
NOTE: The reaction is not set up on ice due to potential precipitation of DNA in the presence of spermidine.
- Butler, E. T., and Chamberline, M. J. (1982) J. Biol. Chem. 257, 5772.
- Kassavetis, G. A., Butler, E. T., Roulland, D. and Chamberlin, M. J. (1982) J. Biol. Chem. 257, 5779.