Natural and electroporation-mediated transformation of Methanococcus voltae protoplasts

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TypeArticle
Journal titleApplied and Environmental Microbiology
Volume60
Pages903907; # of pages: 5
AbstractThe lack of high-efficiency transformation systems has severely impeded genetic research on methanogenic members of the kingdom Archaeobacteria. By using protoplasts of Methanococcus voltae and an integration vector, Mip1, previously shown to impart puromycin resistance, we obtained natural transformation frequencies that were about 80-fold higher (705 transformants per μg of transforming DNA) than that reported with whole cells. Electroporation-mediated transformation of M. voltae protoplasts with covalently closed circular Mip1 DNA was possible, but at lower frequencies of ca. 177 transformants per μg of vector DNA. However, a 380-fold improvement (3,417 transformants per μg of DNA) over the frequency of natural transformation with whole cells was achieved by electroporation of protoplasts with linearized DNA. This general approach, of using protoplasts, should allow the transformation of other methanogens, especially those that may be gently converted to protoplasts as a result of their tendency to lyse in hypotonic solutions.
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LanguageEnglish
AffiliationNational Research Council Canada; NRC Institute for Biological Sciences
Peer reviewedNo
NRC numberPATEL1994
37372
NPARC number9358454
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Record identifier819aed8c-1574-4049-9a27-2bb09f242b03
Record created2009-07-10
Record modified2016-05-09
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