NDLI: Establishment of a gene transfer system for Rhodothermus marinus

Content Provider	Springer Nature Link
Author	Bjornsdottir, S. H. Thorbjarnardottir, S. H. Eggertsson, G.
Copyright Year	2004
Abstract	Genetic manipulation of Rhodothermus marinus has been hampered by the lack of a selection system for gene transfer. We report construction of a Rhodothermus/Escherichia coli shuttle plasmid, containing the R. marinus trpB gene, based on pUC18 and the cryptic R. marinus plasmid pRM21. A plasmid-less R. marinus recipient strain was selected on the basis of growth characteristics and absence of restriction activity. The shuttle plasmid, pRM100, was successfully introduced into a TrpB− mutant of the recipient strain using electroporation and was found to transform it to prototrophy. No loss or rearrangement of pRM100 was observed after growth for 80 generations in non-selective medium. The relative copy numbers of pRM100 and of the parental plasmid, pRM21, were determined as 7±1 and 42±4, respectively. The shuttle plasmid was used to optimize an electroporation protocol, and the maximal number of transformants obtained was 4.3±0.7×106 cfu/μg DNA at 22.5 kV/cm, 200 Ω and 25 μF. Transformation failed, however, after chemical preparation of cells according to several protocols. This is the first report of genetic transformation in the genus Rhodothermus.
Starting Page	675
Ending Page	682
Page Count	8
File Format	PDF
ISSN	01757598
Journal	Applied Microbiology and Biotechnology
Volume Number	66
Issue Number	6
e-ISSN	14320614
Language	English
Publisher	Springer-Verlag
Publisher Date	2004-09-04
Publisher Place	Berlin, Heidelberg
Access Restriction	Subscribed
Content Type	Text
Resource Type	Article
Subject	Medicine Applied Microbiology and Biotechnology Biotechnology

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