Transformation of Dairy Leuconostoc Using Plasmid Vectors from Bacillis, Escherichia, and Lactococcus Hosts

Wyckoff, H.A.; Sandine, W. E.; Kondo, Jeffery K.

doi:10.3168/jds.s0022-0302(91)78303-3

Cited by 15 publications

(12 citation statements)

References 22 publications

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“…The ORF3 disruption by insertion of stop codons was realized by polymerase chain reaction (PCR) using the following synthetic oligonucleotides: OLG1, 5¢-A AACATAGCAGTTGAATTCTTGTTGAG-3¢ (co-ordinates 1691 ±1717) and OLG2, 5¢-TTGAATTCTTTACTTTCAAACG CCC-3¢ [instead of the wild-type sequence 5¢-TTGAATTCTTTGC TTTGAAACGCCC-3¢ (co-ordinates 3285±3261) (underlined nucleotides correspond to an EcoRI site and the sequence alteration is in italics)] and GeneAmp PCR Kit (Perkin-Elmer) as recommended by the supplier. E. coli and JH2-2 strains were transformed by electroporation with a Gene Pulser apparatus (BioRad Laboratories) as described by Dower et al (1988) and Wycko et al (1991), respectively.…”

Section: Dna Manipulations and Genetic Transformationsmentioning

confidence: 99%

Molecular analysis of the replication region of the theta-replicating plasmid pUCL287 from Tetragenococcus (Pediococcus) halophilus ATCC33315

et al. 1997

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The complete nucleotide sequence of the 8.7-kb theta-replicating plasmid pUCL287 from Tetragenococcus halophilus (formerly Pediococcus halophilus) ATCC33315 has been determined. The replication region was identified and analyzed. Its nucleotide sequence contains an untranslated region, the replication origin, followed by two open reading frames (ORFs) encoding two proteins of 311 (RepA287) and 168 (RepB287) amino acids, respectively. Evidence is presented to show that RepA287 represents the plasmid replication protein. RepB287, which is non-essential for replication, is involved in the plasmid copy-number control and segregational stability. The roles of lactococcal proteins homologous to RepB287 have not been defined so far. Nevertheless, the structural organization of the pUCL287 replication region is remarkably similar to those of well known theta-replicating lactococcal plasmids despite the absence of homology of the replication origin and of the replication protein, and this suggests that pUCL287 uses the same mechanism of replication. Nucleotide sequence comparisons show that pSMB74, a pediococcal plasmid encoding bacteriocin production, is a member of the pUCL287 replicon family.

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Section: Dna Manipulations and Genetic Transformationsmentioning

confidence: 99%

Molecular analysis of the replication region of the theta-replicating plasmid pUCL287 from Tetragenococcus (Pediococcus) halophilus ATCC33315

et al. 1997

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“…It gave rise to the pEF17 (' mpoR '), pEF16 (' mptR '), pEF23 (' mphR '), pEF5 (' lpoR '), pEF18 (' mptB ') and pEF19 (' mptD ') plasmids. They were then used to transform E. faecalis JH2-2 as described by Wyckoff et al (1991) to achieve independent gene knockout by homologous recombination with the E. faecalis JH2-2 chromosome. The resulting interrupted mutants (erythromycin-resistant) from each experiment were analysed by Southern blotting of chromosomal DNA (Sambrook et al, 1989), previously digested by HindIII.…”

Section: Methodsmentioning

confidence: 99%

Analysis of σ54-dependent genes in Enterococcus faecalis: a mannose PTS permease (EIIMan) is involved in sensitivity to a bacteriocin, mesentericin Y105

et al. 2001

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, strikingly led to resistance to mesentericin Y105. Moreover, MptD appears to be a peculiar membrane subunit, bearing an additional domain compared to most known IID subunits. According to these results, EII Man t is clearly involved in susceptibility to mesentericin Y105 and could even be its receptor at the E. faecalis surface. Finally, it is hypothesized that MptD could be responsible for the targeting specificity, via an interaction between its additional domain and mesentericin Y105.

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“…cremoris, L. mesenteroides subsp. dextranicum, and L. lactis (19). According to this protocol, cells are grown to mid-log phase in a medium supplemented with threonine.…”

Section: Resultsmentioning

confidence: 99%

Transformation of Leuconostoc carnosum 4010 and Evidence for Natural Competence of the Organism

Helmark

Hansen

Jelle

et al. 2004

Appl Environ Microbiol

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Plasmid transformation in Leuconostoc carnosum 4010 was analyzed. A successful transformation protocol for L. carnosum was established by modifying an existing protocol for Lactococcus lactis. Several parameters, including the number of generations that the cells had grown at the time of harvest, glycine concentration, the time of incubation for phenotypic expression, and the electrical field strength, were investigated and proved to have influence on the transformation frequency. Electrocompetence was found to be transient and to peak in the early exponential growth phase. Optimized conditions resulted in transformation frequencies of up to 6.7 ؋ 10 5 transformants per microgram of plasmid DNA. A total of five plasmids in L. carnosum were successfully introduced and maintained. Interestingly, we discovered that DNA uptake was of a frequency of 3 ؋ 10 ؊6 to 19 ؋ 10 ؊6 transformants per CFU in the absence of an applied electrical field. We concluded that L. carnosum is naturally competent.Lactic acid bacteria (LAB) have the potential to preserve foods and have been used for this purpose for thousands of years. One important attribute of LAB is their ability to produce several metabolic products like organic acids, fatty acids, hydrogen peroxide, and diacetyl that have antimicrobial effects (4,13,15). In recent years interest has been paid to the ability of LAB to produce antimicrobial compounds called bacteriocins. These compounds have potential as natural substitutes for chemical food preservatives in the production of foods with an enhanced shelf life (3,18).Leuconostoc species are widespread in the natural environment and are of technological interest in many different food industries due to their advantageous fermentation characteristics (6). Leuconostoc carnosum is a facultative anaerobic, psychrotrophic LAB, originally associated with spoiled, packaged-meat products (1,8,16). However, investigations have revealed that this spoilage organism is a powerful inhibitor of bacterial growth due to its production of bacteriocins, with particularly strong antagonistic activity against the food-borne pathogen Listeria monocytogenes (2).Advances in genetic technologies have made it possible to develop LAB starter cultures with enhanced fermentation characteristics, and today the genetic manipulation of LAB through the introduction of plasmid DNA has many potential applications in the development of improved food products. Electroporation has been used to transfer DNA into a variety of different LAB as an easy and rapid method compared to most other methods for the transfer of plasmid DNA to bacteria. Protocols for the electroporation of Leuconostoc have been described (5, 12), and in 1991 these protocols were modified by Wycloff et al. (19) for the electroporation of Leuconostoc mesenteroides subsp. cremoris, L. mesenteroides subsp. dextranicum, and Leuconostoc lactis. However, until now, no protocol has been described for L. carnosum.The aim of the present study was to develop a DNA transformation protocol for L. ca...

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Transformation of Dairy Leuconostoc Using Plasmid Vectors from Bacillis, Escherichia, and Lactococcus Hosts

Cited by 15 publications

References 22 publications

Molecular analysis of the replication region of the theta-replicating plasmid pUCL287 from Tetragenococcus (Pediococcus) halophilus ATCC33315

Molecular analysis of the replication region of the theta-replicating plasmid pUCL287 from Tetragenococcus (Pediococcus) halophilus ATCC33315

Analysis of σ54-dependent genes in Enterococcus faecalis: a mannose PTS permease (EIIMan) is involved in sensitivity to a bacteriocin, mesentericin Y105

Transformation of Leuconostoc carnosum 4010 and Evidence for Natural Competence of the Organism

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