HiScribe™ T7 High Yield RNA Synthesis Kit

 

  • Up to 180 μg of RNA per reaction from 1 μg of control template
  • Enables full substitution of NTPs for labeling and incorporation of modified bases
  • Template removal and RNA purification reagents included
  • Linearized control template included for verification of RNA synthesis

Ordering Information

E2040S_pdp_800X450
  • 50 reactions
    $217.00
  • Product Information
    The HiScribe T7 High Yield RNA Synthesis Kit is an extremely flexible system for in vitro transcription of RNA using T7 RNA Polymerase. The kit allows for synthesis many kinds of RNA including internally labeled and co-transcriptionally capped transcripts.

    RNA synthesized from the kit is suitable for many applications including RNA structure and function studies, ribozyme biochemistry, probes for RNase protection assays and hybridization based blots, anti-sense RNA and RNAi experiments, microarray analysis, microinjection, and in vitro translation and RNA vaccines.

    The kit contains sufficient reagents for 50 reactions of 20 μl each. Each standard reaction yields up to 180 μg of RNA from 1 μg control template. Each kit can yield up to 9 mg RNA. For 32P labeling, the kit contains enough reagents for 100 reactions of 20 μl each. 

    Materials Not Included:
    • DNA Template: The DNA template must be linear and contain the T7 RNA Polymerase promoter with correct orientation in relation to target sequence to be transcribed. 
    • Cap Analogs: NEB #S1411, #S1404, #S1405, #S1406 and #S1407 
    • Modified-NTP: Biotin-, Fluorescein-, Digoxigenin-, or Aminoallyl-NTP 
    • Labeling: [α-32P] labeled ribonucleotide (800-6,000 Ci/mmol) 
    • General: 37°C incubator or PCR machine, nuclease-free water 
    • DNase I: DNase I (RNase-free) (NEB #M0303) 
    • Purification: Buffer- or water-saturated phenol/chloroform, ethanol and 3 M sodium acetate, pH 5.2, spin columns 
    • Gel Analysis: Gels and running buffers, gel apparatus, power supply

    Figure 1. Transcription by T7 RNA Polymerase Figure 1. Transcription by T7 RNA Polymerase

    Figure 2. Time course of standard RNA synthesis from three DNA templates Figure 2. Time course of standard RNA synthesis from three DNA templates

    Reactions were incubated at 37°C in a PCR machine. Transcripts were purified by spin columns and quantified on NanoDrop™ Spectrophotometer.
    Figure 3. Effect of template amount on RNA yield Figure 3. Effect of template amount on RNA yield

    Standard reactions were incubated at 37°Cin a PCR machine for 2 hours. Transcripts were purified by spin columns and quantified on NanoDrop™ Spectrophotometer.
    Figure 4: Improved RNA yield and integrity from extended duration transcription reactions Figure 4: Improved RNA yield and integrity from extended duration transcription reactions

    reactions were assembled, in duplicate, according to the manufacturers’ suggested protocols using 3 ng of dsDNA template encoding a 1.8 kb RNA, and incubated at 37°C for 16, 24 and 40 hours. At each time point, the corresponding tubes were transferred to -20°C to stop the reaction. Transcription reactions were column purified after the last time point.

    (A) Transcript yield – After column purification, RNA concentration was measured using a NanoDrop spectrophotometer and total RNA yield was calculated. These data demonstrate that a substantially higher yield of RNA was synthesized using the HiScribe T7 High Yield RNA Synthesis Kit as compared to the competitor’s kit.

    (B) Transcript integrity – 150 ng of column purified RNA was run a 1.2% denaturing agarose gel, stained with ethidium bromide and visualized by UV fluorescence. The data demonstrate greatly improved transcript integrity after extended duration RNA synthesis reactions using the HiScribe T7 High Yield RNA Synthesis Kit as compared to the competitor’s kit.

    Kit Components

    The following reagents are supplied with this product:

    Store at (°C) Concentration
    10X T7 Reaction Buffer -20 10 X
    T7 RNA Polymerase Mix -20
    FLuc Control Template 0.5 μg/μl
    ATP  -20 100 mM
    GTP  -20 100 mM
    UTP  -20 100 mM
    CTP  -20 100 mM
    Product Categories:
    RNA Synthesis Products
    Applications:
    RNA Probe Generation,
    RNA Structural Analysis,
    In vitro Synthesis & Processing
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