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Biotechnology and Bioprocess Engineering

, Volume 19, Issue 2, pp 350–362 | Cite as

A modified piggybac transposon system mediated by exogenous mRNA to perform gene delivery in bovine mammary epithelial cells

  • Guangdong Hu
  • Jing Wang
  • Hui Huang
  • Fusheng Quan
  • Jian Kang
  • Yongyan Wu
  • Yuanpeng Gao
  • Feng Su
  • Minghao Shao
  • Yong ZhangEmail author
Research Paper

Abstract

Transposons are widely used for genetic engineering in various model organisms. Recently, piggyBac (PB) has been developed as a transposable and efficient gene transfer tool in mammalian cells. In the present study, we developed three types of PB transposon systems containing a dual plasmid system (DPS), a single plasmid system (SPS), and a DNA-mRNA combined system (DRPS) and characterized their basic properties in HEK293 cells. The basic elements of the donor plasmid included a selectable-reporter gene expression cassette, two loxP sites in the same orientation, a multiple cloning site, and two chicken β-globin insulator core elements. We further identified the function of the selectable-reporter and examined PB integration sites in the human genome. Moreover, we compared the transposition efficacy and found that SPS transposed more efficiently, as compared to DPS; integration into the host genome was determined by measuring PBase activity. Results discovered the loss of PBase activity in the DRPS, indicating that this system is much more biologically safe, as compared to DPS and SPS. Finally, we employed the DRPS to successfully perform a gene delivery into bovine mammary epithelial cells (BMECs). Taken together, the information from this study will improve the flexibility of PB transposon systems and reduce the genotoxicity of PBase in genetic engineering.

Keywords

piggyBac transposon transposase gene delivery bovine mammary epithelial cells 

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Copyright information

© The Korean Society for Biotechnology and Bioengineering and Springer-Verlag Berlin Heidelberg 2014

Authors and Affiliations

  • Guangdong Hu
    • 1
    • 2
  • Jing Wang
    • 1
  • Hui Huang
    • 1
    • 2
  • Fusheng Quan
    • 1
    • 2
  • Jian Kang
    • 1
    • 2
  • Yongyan Wu
    • 1
    • 2
  • Yuanpeng Gao
    • 1
    • 2
  • Feng Su
    • 1
    • 2
  • Minghao Shao
    • 1
    • 2
  • Yong Zhang
    • 1
    • 2
    Email author
  1. 1.College of Veterinary MedicineNorthwest A&F UniversityYanglingChina
  2. 2.Key Laboratory of Animal Biotechnology, Ministry of AgricultureNorthwest A&F UniversityYanglingChina

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