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T7 P&L RNA Polymerase HC

Highly concentrated and modified T7 RNA Polymerase for set-up of high yield in vitro transcription reactions

Cat. No. Amount Price (EUR) Buy / Note
RNT-018 50 μl (2 μg/μl) 365,00 Add to Basket/Quote Add to Notepad

For in vitro use only!

Shipping: shipped on blue ice

Storage Conditions: store at -20 °C
avoid freeze/thaw cycles

Shelf Life: 12 months

Purity: ≥ 95 % (SDS-PAGE)

Form: liquid

Concentration: 2.0 – 2.2 μg/μl (A280, ɛ = 140 L mmol-1 cm-1 [1] in T7 RNA Polymerase HC Storage Buffer)

Description:
Bacteriophage T7 RNA Polymerase is a DNA-dependent RNA polymerase (99 kDa) that catalyzes in vitro RNA synthesis from a DNA sequence containing a T7 phage promoter.

The highly concentrated version and modified T7 RNA Polymerase HC is ideally suited for set-up and optimization of high yield in vitro transcription reactions. Proline 266 has been replaced by leucine (P266L) which has been associated with

  • decreased abortive transcription[1].
  • increased 5' homogeneity of transcripts synthesized from A-initiating phi2.5 promoter[2]
  • increased 5' incorporation efficiency of GTP analogs[3]
The provided T7 IVT Set-up Buffer, MgCl2 and DTT solutions enable testing of different reaction conditions such as nucleotide/ MgCl2 concentrations. Recommendation for assay set-up with minimal optimization: HighYield T7 P&L RNA Synthesis Kit (#RNT-201).

Activity: 1 μl T7 P&L RNA Polymerase HC generates >/= 30 μg RNA in 10 min at 37°. This is comparable to 1000 U of NEB’s T7 RNA Polymerase (High Concentration), #M0460T (activity assay conditions: 1x T7 IVT Set-up Buffer supplemented with 10 mM DTT, 12 mM MgCl2, 3 mM NTPs, 1 μl T7 RNA Polymerase HC and 1 μg DNA template (1.4 kpb RNA transcript) in 20 μl. RNA yield was determined with a fluorescence microplate assay.)

Content:
T7 P&L RNA Polymerase HC
1x 50 μl (2 μg/μl)
50 mM Tris-HCl, 1 mM EDTA, 100 mM NaCl, 5 mM DTT, 0.1 % Triton-X-100, 50 % Glycerol (v/v), pH 8.0

T7 IVT Set-up Buffer
1x 200 μl (10x)
400 mM Tris-HCl (pH 8.0), 20 mM Spermidin

DTT
1x 100 μl (100 mM)

MgCl2
1x 200 μl (200 mM)

Related products:

Selected References:
[1] Guillerez et al. (2005) A mutation in T7 RNA polymerase that facilitates promoter clearance. Natl. Acad. Sci. U.S.A102:5958.