Source: Purified from an E. coli strain carrying a plasmid with the cDNA o RNA polymerase of T7 phage.

Description: MINOTECH T7 RNA polymerase is an efficient RNA polymerase able to transcribe DNA sequences under the T7 phage promoter up to (5.5 kbs).

Unit definition: One unit is defined as the amount of enzyme that will incorporate 1 nmol ATP into acid-insoluble material in a total reaction volume of 50 μl in 1 hour at 37°C.

Quality Control Assays:

• Functional Assay: T7 RNA polymerase has been tested on PCR products, annealed oligos and digested or supercoiled plasmid DNA containing T7 promoter Sequence: TAATACGACTCACTATA. 100 units of enzyme incubated with the various substrates obtaining transcripts ranging from 120-5500 nucleotides. The full length expected transcripts were found to be above 95% pure based on an ethidium bromide-stained agarose gel.
• Absence of contaminants: Tested extensively for the absence of endo-, exodeoxyribonucleases and RNases.

Guaranteed stability: T7 RNA polymerase is guaranteed to maintain stability until expiration date.

 *PCR products/annealed oligos or plasmid DNA containing T7 promoter

• Incubate at 37^οC for 2 hours.
• Optionally add 0,5 -1 u DNAse I and incubate for 15 min at 37^οC to remove input DNA.
• Purify RNAs by standard phenol chloroform protocol described in Maniatis et al., 1982.

Maniatis, T., E. F. Fritsch, and J. Sambrook. 1982. Molecular cloning: a laboratory manual, p. 194-197. Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y