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Molecular and General Genetics MGG

, Volume 234, Issue 2, pp 265–274 | Cite as

Structural and functional analysis of two cryptic plasmids from Lactobacillus pentosus MD353 and Lactobacillus plantarum ATCC 8014

  • Rob J. Leer
  • Nicole van Luijk
  • Mark Posno
  • Peter H. Pouwels
Article

Summary

The DNA sequences of a 2.4 kb plasmid (p353-2) from Lactobacillus pentosus MD353 and a 1.9 kb plasmid (p8014-2) from Lactobacillus plantarum ATCC 8014 show 81.5% overall similarity. Both plasmids carry elements (replication protein gene, plus-origin and minus-origin of replication), which are typical of plasmids that replicate via a rolling-circle mechanism of replication (RCR). Direct evidence for an RCR mechanism was obtained by showing the accumulation of single-stranded plasmid intermediates in the presence of rifampicin. A minus-origin of replication was defined for plasmids p353-2 and p8014-2 based on DNA sequence analysis and on its ability to convert single-stranded into double-stranded plasmid DNA. Plasmids pLPE323, pLPE350 and pLPC37 that are derived from the p353-2 or p8014-2 replicon are structurally and segregationally stable in L. pentosus MD353, L. plantarum ATCC 8014 and in Lactobacillus casei ATCC 393. The presence of Escherichia coli or λ DNA fragments in vectors derived from p353-2 or p8014-2 does not affect the structural stability but results in segregational instability of the vectors. The instability increases with increasing size of the inserted DNA fragment. Since vectors based on these replicons can be efficiently propagated in a wide variety of Lactobacillus species, they are highly suitable for cloning and expression of foreign DNA in Lactobacillus, provided that selective pressure is applied.

Key words

Lactobacillus plasmids rolling circle replication segregational stability DNA sequence analysis 

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Copyright information

© Springer-Verlag 1992

Authors and Affiliations

  • Rob J. Leer
    • 1
  • Nicole van Luijk
    • 1
  • Mark Posno
    • 1
  • Peter H. Pouwels
    • 1
  1. 1.TNO Medical Biological LaboratoryRijswijkThe Netherlands

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