Chimeric Mos1 and piggyBac transposases result in site-directed integration

K. J. Maragathavally, J. M. Kaminski, C. J. Coates

Research output: Contribution to journalArticle

53 Scopus citations

Abstract

Genetic transformation systems based on Mos1 and piggyBac transposable elements are used to achieve stable chromosomal integration. However, integration sites are randomly distributed in the genome and transgene expression can be influenced by position effects. We developed a novel technology that utilizes chimeric transposases to direct integration into specific sites on a target DNA molecule. The Gal4 DNA binding domain was fused to the NH2 terminus of the Mos1 and piggyBac transposases and a target plasmid was created that contained upstream activating sequences (UAS), to which the Gal4 DBD binds with high affinity. The transpositional activity of the Gal4-Mos1 transposase was 12.7-fold higher compared to controls where the Gal4-UAS interaction was absent and 96% of the recovered transposition products were identical, with integration occurring at the same TA site. In a parallel experiment, a Gal4-piggyBac transposase resulted in an 11.6-fold increase in transpositional activity compared to controls, with 67% of the integrations occurring at a single TTAA site. This technology has the potential to minimize nonspecific integration events that may result in insertional mutagenesis and reduced fitness. Site-directed integration will be advantageous to the manipulation of genomes, study of gene function, and for the development of gene therapy techniques.

Original languageEnglish (US)
Pages (from-to)E1188-E1195
JournalFASEB Journal
Volume20
Issue number11
DOIs
StatePublished - Sep 2006

Keywords

  • Gal4-UAS
  • Mariner
  • Transposition assay

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry
  • Molecular Biology
  • Genetics

Fingerprint Dive into the research topics of 'Chimeric Mos1 and piggyBac transposases result in site-directed integration'. Together they form a unique fingerprint.

Cite this