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    LITMUS™ U is a linear vector with 8 nucleotide, 3´non-complementary overhangs that can be used for the rapid cloning of PCR fragments without ligation (Figure 1). LITMUS U has opposing T7 promoters which generate dsRNA from a cloned fragment with little additional sequence by in vitro transcription with T7 RNA Polymerase.

    To clone into LITMUS U, PCR products must be generated using primers that have the following sequences at the 5´ end: 5´ GGAGACAUNNN... and 5´ GGGAAAGUNNN... (U = uracil and NNN... = specific primer sequence). After PCR, the DNA product can be directly cloned into LITMUS U using the USER Enzyme (NEB #M5505 ) in a 30 minute reaction without ligation or any purification steps (1) following the USER Friendly Cloning Kit (NEB #E5500) protocol.

    Figure 1: Cloning a PCR Product into LITMUS U.


    • Fast, simple & extremely efficient cloning of PCR Products 
    • Mix PCR product with USER™ Enzyme and vector, incubate for 30 minutes, transform
    • No PCR product cleanup required

    Properties and Usage

    Storage Temperature


    Quality Control

    Quality Control Assays

    The following Quality Control Tests are performed on each new lot and meet the specifications designated for the product. Individual lot data can be found on the Product Summary Sheet/Datacard or Manual which can be found in the Supporting Documents section of this page.
    • Functional Test (USER Enzyme):
      The USER™ (uracil-specific excision reagent) enzyme is tested for its ability to nick a PCR product at a uracil contained in the primers and assemble with a vector containing complementary overlaps; transformation and plating results in >95% white colonies containing insert.


    1. Bitinaite, J. et al. Unpublished observation

    Supporting Documents

    Material Safety Datasheets

    The following is a list of Material Safety Data Sheets (MSDS) that apply to this product to help you use it safely. The following file naming structure is used to name these document files: [Product Name] MSDS. For international versions please contact us at info@neb.com.


    The Product Summary Sheet, or Data Card, includes details for how to use the product, as well as details of its formulation and quality controls. The following file naming structure is used to name the majority of these document files: [Catalog Number]Datasheet-Lot[Lot Number]. For those product lots not listed below, please contact NEB at info@neb.com or fill out the Technical Support Form for appropriate document.
    1. Cloning with USER Enzyme

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