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Molecular characterization of Bifidobacterium longum biovar longum NAL8 plasmids and construction of a novel replicon screening system.
Appl Microbiol Biotechnol. 2007 Apr; 74(5):1053-61.AM

Abstract

In this study, we performed molecular characterization and sequence analysis of three plasmids from the human intestinal isolate Bifidobacterium longum biovar longum NAL8 and developed a novel vector screening system. Plasmids pNAL8H (10 kb) and pNAL8M (4.9 kb) show close sequence similarity to and the same gene organization as the already characterized B. longum plasmids. The B. longum plasmid pNAC1 was identified as being most closely related to pNAL8L (3.5 kb). However, DNA sequence analysis suggested that direct repeat-rich sites could have promoted several recombination events to diversify the two plasmid molecules. We verified the likely rolling circle replication of plasmid pNAL8L and studied the phylogenetic relationship in all the Bifidobacterium plasmids fully sequenced to date based on in silico comparative sequence analysis of their replication proteins and iteron regions. Our transformation experiments confirmed that the ColE1 replication origin from high-copy-number pUC vectors could interfere with the replication apparatus of Bifidobacterium plasmids and give rise to false positive clones. As a result, we developed a system suitable for avoiding possible interference by other functional replication modules on the vector and for screening functional replicons from wild-type plasmids.

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Authors+Show Affiliations

Guglielmetti S
Department of Food Science and Microbiology, Industrial Microbiology Section, University of Milan, Via Celoria 2, 20133 Milan, Italy. simone.guglielmetti@unimi.it
Karp M
No affiliation info available
Mora D
No affiliation info available
Tamagnini I
No affiliation info available
Parini C
No affiliation info available

MeSH

BifidobacteriumCloning, MolecularGenetic VectorsPhylogenyPlasmidsReplicon

Pub Type(s)

Journal Article

Language

eng

PubMed ID

17151871

Citation

Guglielmetti, Simone, et al. "Molecular Characterization of Bifidobacterium Longum Biovar Longum NAL8 Plasmids and Construction of a Novel Replicon Screening System." Applied Microbiology and Biotechnology, vol. 74, no. 5, 2007, pp. 1053-61.
Guglielmetti S, Karp M, Mora D, et al. Molecular characterization of Bifidobacterium longum biovar longum NAL8 plasmids and construction of a novel replicon screening system. Appl Microbiol Biotechnol. 2007;74(5):1053-61.
Guglielmetti, S., Karp, M., Mora, D., Tamagnini, I., & Parini, C. (2007). Molecular characterization of Bifidobacterium longum biovar longum NAL8 plasmids and construction of a novel replicon screening system. Applied Microbiology and Biotechnology, 74(5), 1053-61.
Guglielmetti S, et al. Molecular Characterization of Bifidobacterium Longum Biovar Longum NAL8 Plasmids and Construction of a Novel Replicon Screening System. Appl Microbiol Biotechnol. 2007;74(5):1053-61. PubMed PMID: 17151871.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Molecular characterization of Bifidobacterium longum biovar longum NAL8 plasmids and construction of a novel replicon screening system. AU - Guglielmetti,Simone, AU - Karp,Matti, AU - Mora,Diego, AU - Tamagnini,Isabella, AU - Parini,Carlo, Y1 - 2006/12/07/ PY - 2006/10/09/received PY - 2006/11/09/accepted PY - 2006/11/08/revised PY - 2006/12/8/pubmed PY - 2007/9/29/medline PY - 2006/12/8/entrez SP - 1053 EP - 61 JF - Applied microbiology and biotechnology JO - Appl Microbiol Biotechnol VL - 74 IS - 5 N2 - In this study, we performed molecular characterization and sequence analysis of three plasmids from the human intestinal isolate Bifidobacterium longum biovar longum NAL8 and developed a novel vector screening system. Plasmids pNAL8H (10 kb) and pNAL8M (4.9 kb) show close sequence similarity to and the same gene organization as the already characterized B. longum plasmids. The B. longum plasmid pNAC1 was identified as being most closely related to pNAL8L (3.5 kb). However, DNA sequence analysis suggested that direct repeat-rich sites could have promoted several recombination events to diversify the two plasmid molecules. We verified the likely rolling circle replication of plasmid pNAL8L and studied the phylogenetic relationship in all the Bifidobacterium plasmids fully sequenced to date based on in silico comparative sequence analysis of their replication proteins and iteron regions. Our transformation experiments confirmed that the ColE1 replication origin from high-copy-number pUC vectors could interfere with the replication apparatus of Bifidobacterium plasmids and give rise to false positive clones. As a result, we developed a system suitable for avoiding possible interference by other functional replication modules on the vector and for screening functional replicons from wild-type plasmids. SN - 0175-7598 UR - https://www.unboundmedicine.com/medline/citation/17151871/Molecular_characterization_of_Bifidobacterium_longum_biovar_longum_NAL8_plasmids_and_construction_of_a_novel_replicon_screening_system_ L2 - https://dx.doi.org/10.1007/s00253-006-0755-1 DB - PRIME DP - Unbound Medicine ER -