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Pharmaceutical Research

, Volume 29, Issue 4, pp 1040–1045 | Cite as

Nonreplicating Intracellular Bacterial Vector for Conjugative DNA Transfer into Mitochondria

  • Young Geol Yoon
  • Michael D. Koob
Research Paper

Abstract

Purpose

We have previously shown that DNA constructs can be introduced into isolated mitochondria through the process of conjugative transfer from an E. coli host. We set out to generate a conjugative E. coli strain that would be able to introduce itself into the cytoplasm of a mammalian cell for the purpose of transferring DNA into the mitochondria in the cell.

Methods

We have now developed a method for making E. coli strains from which nonreplicating populations of daughter cells can be generated. We used this approach to modify a facultative intracellular enteroinvasive E. coli (EIEC) and introduced conjugative functions to this new strain.

Results

We demonstrate that this new strain can generate large populations of nonreplicating cells that are capable of conjugative transfer to other cells and can readily invade mammalian tissue culture cells, live in the cytoplasm of the cell for several days, and that do not kill the invaded mammalian cell.

Conclusions

We successfully constructed an E. coli host suitable for intracellular conjugative transfer but, due to the lack of suitable mitochondrial screening or selectable markers, we have not yet been able to determine if these bacterial vectors can in fact transfer DNA into intracelluar mitochondria.

KEY WORDS

conjugative DNA transfer facultative intracellular enteroinvasive Escherichia coli mitochondria mitochondrial DNA nonreplicating bacterial cell 

Abbreviations

eGFP

enhanced green fluorescent protein

EIEC

enteroinvasive Escherichia coli

PCR

Polymerase Chain Reaction

Notes

Acknowledgments & Disclosures

We thank Michael Sadowsky for the gifts of plasmids and E.coli strains. This work was funded by the National Institutes of Health (NS052612) and by the Minnesota Partnership for Biotechnology and Medical Genomics.

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Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  1. 1.Laboratory Medicine and PathologyUniversity of MinnesotaMinneapolisUSA
  2. 2.Department of Biomedical SciencesJungwon UniversityChungcheongbuk-doKorea

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