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v.2 MuA Transposase (in vitro Manipulator)

Domus v.2 MuA Transposase (in vitro Manipulator) is a hyperactive MuA transposase variant (1). It has a superior performance even in the most demanding in vitro applications, with a reaction time of only 10 minutes. When v.2 MuA Transposase is incubated in vitro with Mu transposon, stable transpososomes are formed. Under reaction conditions with Mg2+, the transpososome becomes activated and executes transposon integration into the target DNA. Transposons can contain any DNA (e.g. selectable markers or other genetic elements), between MuA binding sites at both ends.

Catalog Number Unit Size Price   
D010201 2.2 µg (0.22 µg/µl) 106 € Buy now
D010201M 4.4 µg (0.22 µg/µl) 191 € Buy now
D010201L 11 µg (0.22 µg/µl) 485 € Buy now

v.2 MuA outperforms wild-type MuA with modified transposon ends

Feature v.2 MuA Wild‑type MuA
Mu end type Modified Mu ends (e.g., BpuEI site) Modified Mu ends (e.g., BpuEI site)
Reaction time 10 min 60 min
Integrants (CFU per µg transposon DNA) 1.1 × 10⁶ 9.8 × 10⁴
Overall performance Fast, high yield standard yield

Using Mu ends modified with a BpuEI restriction site, v.2 MuA produced 1.1 × 10⁶ CFU per µg of transposon DNA in a 10-minute reaction, whereas wild-type MuA generated 9.8 × 10⁴ CFU per µg after a 60-minute incubation under standard reaction conditions.

These results demonstrate that v.2 MuA is both faster and more efficient than wild-type MuA when working with modified transposon ends, making it a strong choice for demanding cloning and library-generation applications.

Applications

  • In vitro insertion of Mu transposon into linear DNA or plasmids
  • In vitro insertion of Mu transposon into large genomic clones (e.g. BACs, YACs)

Benefits

  • Hyperactive (1)
  • Short reaction time, only 1-10 minutes (1)
  • Simple in vitro reaction, with only transposon DNA, target DNA, v.2 MuA Transposase and a simple reaction buffer (2, 3)
  • Nearly random insertion profile (4, 5)

Scheme for in vitro transposition integration.  A tetramer of MuA transposase and Mu transposon ends assemble into a stable transpososome. Under reaction conditions with Mg2+, the transpososome becomes activated and executes transposon integration into the target DNA.

Guidance for Large‑Scale Library Generation

For projects requiring large mutant libraries (typically above 1 × 10⁴ variants), a
pooling‑based strategy is recommended. This approach, originally described by
Poussu et al. (2004, 2005), involves performing several parallel transposition reactions and
combining them to achieve the desired library size and diversity.

During downstream processing, standard organic extraction steps are used to dissociate
transpososomes and clean the DNA. Phenol treatment disrupts protein–DNA complexes,
chloroform removes residual phenol, and ethanol precipitation provides a routine method
for concentrating the material.

When scaling up, you may use higher absolute amounts of transposon DNA and MuA to
increase overall output. The key requirement is that the
stoichiometry between transposon and MuA remains constant,
ensuring consistent reaction performance.

 

References

1.Rasila,T.S., Pulkkinen,E., Kiljunen,S., Haapa-Paananen,S., Pajunen,M.I., Salminen,A., Paulin,L., Vihinen,M., Rice,P.A. and Savilahti,H. (2018) Mu transpososome activity-profiling yields hyperactive MuA variants for highly efficient genetic and genome engineering. Nucleic Acids Res., 46, 4649–4661.
2. Haapa S, Taira S, Heikkinen E, Savilahti H (1999) An efficient and accurate integration of mini-Mu transposons in vitro: a general methodology for functional genetic analysis and molecular biology applications. Nucleic Acids Res 27:2777-2784
3. Savilahti H, Rice PA, Mizuuchi K (1995) The phage Mu transpososome core: DNA requirements for assembly and function. EMBO J 14:4893-4903
4. Haapa-Paananen S, Rita H, Savilahti H (2002) DNA transposition of bacteriophage Mu. A quantitative analysis of target site selection in vitro. The Journal of Biological Chemistry 277:2843-2851
5. Mizuuchi M, Mizuuchi K (1993) Target site selection in transposition of phage Mu. Cold Spring Harb Symp Quant Biol 58:515-523

Patent
v.2 MuA Transposase (in vitro Manipulator) is covered by International Patent No. WO 2014/013127 Al

 For Research Use Only. Not for use in diagnostic procedures.