Applications of In Vitro Transposomics™

The Applications of Transposomics™ Are Limited Only by Our Imagination

The field of Transposomics™ exploits the ability of certain transposase enzymes to catalyze the random "hop" or insertion of an "artificial" transposon into any other DNA during a brief in vitro or in vivo transposition reaction. The transposon can be any DNA that has a transposase recognition sequence on each end (e.g., Figure 1). An artificial transposon doesn't encode a transposase, but can encode selectable markers, priming sites, origins of replication, RNA polymerase promoters, or other genetic elements or control sequences.

Figure 1. An EZ-Tn5™ Transposon. An EZ-Tn5 Transposon can be any DNA sequence that is between properly-oriented 19-bp inverted repeat Mosaic End (ME) sequences that are specifically and uniquely recognized by EZ-Tn5™ Transposase. The EZ-Tn5 System is based on the hyperactive Tn5 in vitro transposition system described by IY Goryshin & WS Reznikoff1. This system retains the insertion characteristics of Tn5–the most random transposon system known–but has a transposition frequency 1000-fold higher than wild-type Tn5. Epicentre also has a HyperMu™ In Vitro Transposition System that is 50-100 times more active than other MuA Transposase Systems.

In DNA Sequencing, we presented a comparison of the properties of transposon systems and our system for naming transposons. We also described how EZ-Tn5™ or HyperMu™ in vitro transposition systems can be used to simplify and speed up DNA sequencing. In following categories, we discuss how In Vivo Transposomics is revolutionizing microbial genetics by providing tools for studying organisms for which genetic tools were not previously available, and methods and new transposon tools for protein engineering and mapping of protein domains or epitopes.

Even the methods and products covered in other categories do not cover all of the applications or indicate the full power of Transposomics, nor could we hope to do so in this catalog. Our goal in the present category is to provide a few examples of additional applications and transposon tools for In Vitro Transposomics. The rest is up to your imagination.

Examples of In Vitro Transposomics Applications

Although the above examples are given for EZ-Tn5 Systems, researchers can develop similar transposons for use in vitro with Epicentre's HyperMu™ MuA Transposase. The first in vitro transposition system was developed by Dr. Kiyoshi Mizuuchi using the well-characterized temperate bacteriophage Mu2,3. However, until the introduction of Epicentre's HyperMu™ MuA Transposase, Mu-based in vitro transposition systems have not had transposition efficiencies that approached those of Epicentre's EZ-Tn5 Insertion Systems. High transposition efficiencies are critical for some applications. Epicentre's HyperMu MuA Transposase is a hyperactive enzyme that retains the random insertion characteristics of wild-type MuA transposase4 but is at least 50 times more active in vitro than the MuA transposase available from other suppliers. Also, since HyperMu MuA Transposase recognizes the same R1 and R2 end sequences as wild-type MuA Transposase5 , it can be used with HyperMu transposons as well as other artificial Mu transposons.

Strategies That Use Both EZ-Tn5 and HyperMu Transposon Systems

EZ-Tn5 and HyperMu Transposases do not recognize the same end sequences for transposition. Therefore, both systems can be used in strategies in which it is desirable to use more than one transposon system.

References

  1. IY Goryshin & WS Reznikoff (1998) J Biol Chem 273, 7367.
  2. Mizuuchi, K (1983) Cell 35, 785.
  3. Chaconas, G & Harshey, RM (2002) in: Mobile DNA II, ASM Press, Washington, D.C.
  4. Butterfield, YSN et al.(2002) Nucleic Acids Res. 30, 2460.
  5. Savilahti, H et al. (1995) EMBO J. 14, 4893.

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