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Nucleic Acids Research 2005 33(16):e139; doi:10.1093/nar/gni140
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Published online 12 September 2005

© The Author 2005. Published by Oxford University Press. All rights reserved
The online version of this article has been published under an open access model. Users are entitled to use, reproduce, disseminate, or display the open access version of this article for non-commercial purposes provided that: the original authorship is properly and fully attributed; the Journal and Oxford University Press are attributed as the original place of publication with the correct citation details given; if an article is subsequently reproduced or disseminated not in its entirety but only in part or as a derivative work this must be clearly indicated. For commercial re-use, please contact journals.permissions{at}oxfordjournals.org

Methods Online

Transformation of isolated mammalian mitochondria by bacterial conjugation

Young Geol Yoon1,2 and Michael D. Koob1,2,*

1Department of Laboratory Medicine and Pathology, University of Minnesota 420 Delaware Street SE, Minneapolis, MN 55455, USA 2Institute of Human Genetics, University of Minnesota 420 Delaware Street SE, Minneapolis, MN 55455, USA

*To whom correspondence should be addressed. Tel: +1 612 626 6516; Fax: +1 612 626 7031; Email: koobx001{at}umn.edu

Received July 22, 2005. Revised August 18, 2005. Accepted August 26, 2005.

We have developed a method for transferring exogenous DNA molecules into isolated mammalian mitochondria using bacterial conjugation. In general, we accomplish this by (i) inserting an origin of DNA transfer (oriT) sequence into a DNA construct, (ii) transforming the construct into an appropriate Escherichia coli strain and then (iii) introducing the mobilizable DNA into mitochondria through conjugation. We tested this approach by transferring plasmid DNA containing a T7 promoter sequence into mitochondria that we had engineered to contain T7 RNA polymerase. After conjugation between E.coli and mitochondria, we detected robust levels of T7 transcription from the DNA constructs that had been transferred into the mitochondria. This approach for engineering DNA constructs in vitro and subsequent transfer into mitochondria by conjugation offers an attractive experimental system for studying many aspects of vertebrate mitochondrial gene expression and is a potential route for transforming mitochondrial networks within mammalian cells.


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