PiggyBac transposase (PBase)

Versatile DNA transposase from Trichoplusia ni cabbage looper moth that mediates precise cut-and-paste transposition through TTAA target sites. Achieves 50-fold higher genomic integration efficiency than random integration in mammalian cells, enabling applications in gene therapy, CAR-T cell engineering, and iPSC generation.

Length: 1785 bp(595 aa)

Recognition site: TTAA (4 bp)

Directionality: Bidirectional

Efficiency: >50-fold enhancement over random integration in mammalian cells; 34.8% transgenic founder rate in mice with helper plasmid co-injection

Origin: Trichoplusia ni

Characteristics

Catalyzes precise DNA excision and integration at tetranucleotide TTAA target sites, which are duplicated upon insertion. Functions bidirectionally to mediate both integration and precise excision from genomic loci without leaving footprints. Recognizes 13 bp terminal inverted repeats flanking transposon cargo. The 594 amino acid protein achieves robust transposition across diverse mammalian cell types including ES cells, primary T cells, and fibroblasts. Preferentially integrates into transcription units (67% of insertions) with minimal chromosomal regional bias. Maintains stable transgene expression through multiple cell divisions and germline transmission in mice.

Applications: Non-viral gene delivery vector for gene therapy applications including CAR-T cell manufacturing for cancer immunotherapy and correction of genetic diseases. Generates transgenic cell lines and animal models through efficient germline transmission. Enables iPSC reprogramming and stem cell engineering with reduced immunogenicity compared to viral vectors. Clinical applications include EGFR-targeted CAR-T cells for non-small cell lung cancer treatment. Supports large cargo capacity insertional mutagenesis screens for gene discovery.

Limitations: Requires co-delivery of transposase-encoding plasmid or mRNA, adding complexity to vector design. Persistent transposase expression may cause unwanted remobilization of integrated elements, requiring temporal control mechanisms. TTAA target site preference may limit insertion site diversity compared to some viral vectors. Integration into transcription units (while beneficial for expression) increases theoretical insertional mutagenesis risk. Native transposase may exhibit sub-optimal activity in mammalian systems without codon optimization or hyperactive variants.

Sequence

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References

  1. Fraser et al. (1996). Precise excision of TTAA-specific lepidopteran transposons piggyBac (IFP2) and tagalong (TFP3) from the baculovirus genome in cell lines from two species of Lepidoptera. Insect Molecular Biology - Fraser 1996 PiggyBac excision
  2. Zhang et al. (2021). Phase I clinical trial of EGFR-specific CAR-T cells generated by the piggyBac transposon system in advanced relapsed/refractory non-small cell lung cancer patients. Journal of Cancer Research and Clinical Oncology - Zhang 2021 PiggyBac CAR-T