Enzymes for Molecular Biology: Enzymes for Molecular Biology

T7 R&DNA™ Polymerase* is a recombinant mutant form of T7 RNA polymerase (Y639F mutant), in which Tyr-639 has been replaced by Phe.^1,2 This mutation enables T7 R&DNA Polymerase to incorporate 2´-deoxyribonucleoside triphosphates (dNTPs) into full-length transcripts much more efficiently than the corresponding wild-type enzyme,¹ while retaining the same catalytic activity for incorporation of canonical NTPs, and the same high promoter specificity as wild-type T7 RNA Polymerase. The ability of this mutant enzyme to incorporate certain 2´-modified-dNTPs (including but not limited to dNTPs having a 2´-fluorine or 2´-amino group) enables efficient in vitro synthesis of transcripts composed of mixed rNMP/2´-dNMP or rNMP/2´- modified-dNMP from any DNA template that is downstream of the T7 RNA polymerase promoter. Such modified transcripts are useful for many applications.

Note: Complete substitution of one 2´-dNTP (or one 2´-modified dNTP) for a canonical rNTP in a T7 R&DNA Polymerase reaction results in a slight decrease in yield. Additional substitutions of 2´-dNTPs (or 2´-modified-dNTPs) for canonical rNTPs will further reduce yield of the transcript produced. Substitution with all four 2´-dNTPs (or 2´-modified dNTPs) will result in extremely low yields of transcript and is NOT RECOMMENDED.

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Protocol for: T7 R&DNA™ Polymerases

T7 & SP6 R&DNA™ Polymerase Protocol

(catalogue number D7P9201K / D7P9205K / D6P9301K / D6P9305K)

Please note: all protocols off site are the responsibility of the products supplier

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References

1. Sousa, R and Padilla, R. (1995) EMBO J. 14 (18), 4609.
2. U.S. Patent Nos. 5,849,546 and 6,107,037.

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*T7 R&DNA Polymerase is covered by patents issued and pending.

These products are accompanied by a limited non-exclusive license for the purchaser to use the purchased product(s) solely for life science research. No license for any other field of use or immunity under any patent covering any other use is either granted or implied by the sale of the product. Contact the Intellectual Property Manager at Epicentre® Biotechnologies for information on licenses for uses other than life science research.

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Applications

• Synthesis of "RNA" transcripts of mixed rNMP/2´-dNMP or rNMP/2´-modified-NMP composition.
• Synthesis of modified "RNA" transcripts that are resistant to RNase A-type RNases

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