Characterization of pURB500 from the archaeon Methanococcus maripaludis and construction of a shuttle vector

Debra L. Tumbula, Timothy L. Bowen, William B. Whitman

Research output: Contribution to journalArticlepeer-review

36 Scopus citations

Abstract

The complete sequence of the 8,285-bp plasmid pURB500 from Methanococcus maripaludis C5 was determined. Sequence analysis identified 18 open reading frames as well as two regions of potential iterons and complex secondary structures. The shuttle vector, pDLT44, for M, maripaludis JJ was constructed from the entire pURB500 plasmid and pMEB.2, an Escherichia coli vector containing a methanococcal puromycin-resistance marker (P. Gernhardt, O. Possot, M. Foglino, L. Sibold, and A. Klein, Mol. Gen. Genet. 221:273-279, 1900). By using polyethylene glycol transformation, M. maripaludis JJ was transformed at a frequency of 3.3 x 107 transformants per μg of pDLT44. The shuttle vector was stable in E. coli under ampicillin selection but was maintained at a lower copy number than pMEB.2. Based on the inability of various restriction fragments of pURB500 to support maintenance in M. maripaludis JJ, multiple regions of pURB500 were required. pDLT44 did not replicate in Methanococcus voltae.

Original languageEnglish (US)
Pages (from-to)2976-2986
Number of pages11
JournalJournal of bacteriology
Volume179
Issue number9
DOIs
StatePublished - May 1997
Externally publishedYes

ASJC Scopus subject areas

  • Microbiology
  • Molecular Biology

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