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Applied and Environmental Microbiology, October 2003, p. 6121-6127, Vol. 69, No. 10
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.10.6121-6127.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Marker Rescue Studies of the Transfer of Recombinant DNA to Streptococcus gordonii In Vitro, in Foods and Gnotobiotic Rats

Mitra Kharazmi,1 Silke Sczesny,2 Michael Blaut,2 Walter P. Hammes,1 and Christian Hertel1*

Institute of Food Technology, University of Hohenheim, Stuttgart,1 Department of Gastrointestinal Microbiology, German Institute of Human Nutrition, Potsdam-Rehbrücke, Germany2

Received 7 March 2003/ Accepted 31 July 2003

A plasmid marker rescue system based on restoration of the nptII gene was established in Streptococcus gordonii to study the transfer of bacterial and transgenic plant DNA by transformation. In vitro studies revealed that the marker rescue efficiency depends on the type of donor DNA. Plasmid and chromosomal DNA of bacteria as well as DNA of transgenic potatoes were transferred with efficiencies ranging from 8.1 x 10-6 to 5.8 x 10-7 transformants per nptII gene. Using a 792-bp amplification product of nptII the efficiency was strongly decreased (9.8 x 10-9). In blood sausage, marker rescue using plasmid DNA was detectable (7.9 x 10-10), whereas in milk heat-inactivated horse serum (HHS) had to be added to obtain an efficiency of 2.7 x 10-11. No marker rescue was detected in extracts of transgenic potatoes despite addition of HHS. In vivo transformation of S. gordonii LTH 5597 was studied in monoassociated rats by using plasmid DNA. No marker rescue could be detected in vivo, although transformation was detected in the presence of saliva and fecal samples supplemented with HHS. It was also shown that plasmid DNA persists in rat saliva permitting transformation for up to 6 h of incubation. It is suggested that the lack of marker rescue is due to the absence of competence-stimulating factors such as serum proteins in rat saliva.

* Corresponding author. Mailing address: Institute of Food Technology, University of Hohenheim, Garbenstr. 28, D-70599 Stuttgart, Germany. Phone: 49 711 459 4255. Fax: 49 711 459 4199. E-mail: hertel{at}uni-hohenheim.de.

Applied and Environmental Microbiology, October 2003, p. 6121-6127, Vol. 69, No. 10
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.10.6121-6127.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.





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