Innovative Tools for Molecular and Cell Biology

Innovative Tools for Molecular and Cell Biology

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Lactococcus Expression Systems

Constitutive Gene Expression System for Lactococcus lactis and Other Lactic Acid Bacteria

 Small scale L. lactis culture

Small scale L. lactis culture

Features

  • Less endogenous and no exogenous proteases
  • Endotoxin-free expression system
  • Food grade protein expression possible
  • No formation of inclusion bodies
  • No formation of spores
  • Simple fermentation, scale-up, and downstream processing
  • Suitable also for bacteria other than Lactococcus lactis

Description

  • Next to the wealth of traditional food applications, L. lactis is increasingly used for modern biotechnological applications such as the production of recombinant proteins for food, feed, pharmacological and biocatalysis applications.
  • The Constitutive Gene Expression System for Lactococcus lactis and Other Lactic Acid Bacteria, developed by NIZO food research BV, The Netherlands, is easy-to-operate and has advantages for many applications:
  • Overexpression of homologous and heterologous genes for functional studies and to obtain large quantities of specific gene products
  • Metabolic engineering
  • Suitability for protein secretion (Novotny, R. et al. 2005; Ravn, P. et al. 2003; van Asseldonk et al. 1990; Vos, P. et al. 1989) and anchoring in the cell envelope
  • Large scale applications

Plasmids

Vector map of pNZ2103
  1. enlarge

Vector map of pNZ2103

  • The replicons of the vectors pNZ124, pNZ2105, pNZ2103, pNZ7021, pNZ2122, and pNZ2123 are identical and came originally from the Lactococcus lactis plasmid pSH71. However, this replicon has a broad host range. Plasmids with this replicon can replicate in many Gram-positive bacteria, such as Lactobacillus plantarum and Streptococcus thermophilus.

  • pNZ124 – General broad host range cloning vector with multiple cloning site for L. lactis and other lactic acid bacteria. Chloramphenicol selection. (Platteeuw et al., 1994)

  • pNZ2105 – General food grade cloning vector with multiple cloning site for L. lactis NZ3000. LacF-based selection. (Platteeuw et al., 1996)

  • pNZ2103 – Broad host range vector for constitutive gene expression under control of the lacA promoter. For transcriptional fusion. Chloramphenicol selection. (Platteeuw et al., 1996)

  • pNZ7021 – Broad host range vector for constitutive gene expression under control of the strong pepN promoter. For transcriptional fusion. Chloramphenicol selection. (Wegkamp et al., 2007)

  • pNZ2122Food grade vector for constitutive gene expression under control of the lacA promoter for L. lactis NZ3000. For transcriptional fusions. LacF-based selection. (Platteeuw et al., 1996)

  • pNZ2123Food grade vector. Identical to pNZ2122, but with inverted multiple cloning site (MCS). LacF-based selection. (Platteeuw et al., 1996)

  • pNZ2125Food grade cloning vector with multiple cloning site appropriate for gene expression in L. lactis strain NZ1330; alr-based selection. (Platteeuw et al., 1996)

  • pNZ7025Food grade vector for constitutive gene expression under control of the pepN promoter; alr-based selection. (Wegkamp et al., 2007)

Host Strains

Electron microscope image of Lactococcus lactis

Electron microscope image of Lactococcus lactis

Description

  • Lactococcus lactis subsp. cremoris MG1363
  • Plasmid-free progeny of the dairy strain NCDO712. Most widely used host strain for cloning and gene expression in L. lactis (Gasson, 1983).

  • Lactococcus lactis NZ3000
  • Standard strain for food grade selection based upon the ability to grow on lactose. The lactose operon, that is generally present on plasmids, has been integrated into the chromosome and the lacF gene was deleted. Deletion of the lacF gene makes this strain unable to grow on lactose unless lacF is provided on a plasmid (van Alen-Boerrigter, I. J., and W. M. de Vos, unpublished data; de Ruyter et al., 1996).

  • Lactoccoccus lactis NZ1330
  • This host strain is made for food grade expression systems based on an alr selection marker. It was generated by deleting the alr gene, an alanine racemase encoding gene, in the progenitor strain MG1363. Deletion of the alr gene results in auxotrophy for the essential component D-alanine, implying that the strain is unable to grow on media without D-alanine unless alr is provided on a plasmid, and thereby enables the conversion of L-Ala to D-Ala. The alr gene serves as food grade complementation marker (Bron et al., 2002).
Strains Plasmids Medium
Lactococcus lactis MG1363 non-food grade M17 + 0.5% glucose + 10 µg/ml chloramphenicol
Lactococcus lactis NZ3000 food grade see Handbook (PDF)
Lactococcus lactis NZ1330 food grade see Handbook (PDF)

ORDER INFORMATION

  • For shipping and storage information please click on Order#.
Order# Description Amount Price Data Sheet
VS-ELV01100 pNZ124 Lactococcus lactis cloning vector 10 µg 460,00 PDF
VS-ELV01150 pNZ2105 Lactococcus lactis cloning vector, food grade 10 µg 482,00 PDF
VS-ELV01200 pNZ2103 Lactococcus lactis const. vector, lacA 10 µg 471,00 PDF
VS-ELV01250 pNZ7021 Lactococcus lactis const. vector, pepN 10 µg 471,00 PDF
VS-ELV01300 pNZ2122 Lactococcus lactis const. vector, mcs X-H, food grade 10 µg 493,00 PDF
VS-ELV01350 pNZ2123 Lactococcus lactis const. vector, mcs H-X, food grade 10 µg 493,00 PDF
VS-ELV01400 pNZ2125 Lactococcus lactis const. vector, food grade, (alr) 10 µg 493,00 PDF
VS-ELV01450 pNZ7025 Lactococcus lactis const. vector, pepN, food grade, (alr) 10 µg 493,00 PDF
VS-ELS01363 Lactococcus lactis expression strain MG1363 1 ml 294,00 PDF
VS-ELS03000 Lactococcus lactis expression Strain NZ3000, food grade 1 ml 312,00 PDF
VS-ELS01330-01 NICE Lactococcus lactis expression Strain NZ1330, food grade, (alr) 1 ml 383,00 PDF
VS-ELS10610-01 NICE E. coli host Strain MC1061 1 ml 305,00 PDF
VS-ELS10710-01 MC1061 Chemically Competent Escherichia coli 5 x 100 µl 438,00 PDF
  • All prices are in EURO excl. VAT and shipping. For further pricing and order information please ask your local distributor.

Download

  • Constitutive Expression System for Lactococcus lactis Handbook (PDF)

Vector maps and Sequences

Vector map of pNZ124 (pic)
Vector map of pNZ2105 (pic)
Vector map of pNZ2103 (pic)
Vector map of pNZ7021 (pic)
Vector map of pNZ2122 (pic)
Vector map of pNZ2123 (pic)
Vector map of pNZ2125 (pic)
Vector map of pNZ7025 (pic)

Sequence of pNZ124 (txt)
Sequence of pNZ2105 (txt)
Sequence of pNZ2103 (txt)
Sequence of pNZ7021 (txt)
Sequence of pNZ2122 (txt)
Sequence of pNZ2123 (txt)
Sequence of pNZ2125 (txt)
Sequence of pNZ7025 (txt)

Literature

  • de Ruyter, P. G., Kuipers, O.P. and W. M. de Vos. ( 1996 b). Controlled gene expression systems for Lactococcus lactis with the food grade inducer nisin. Applied and Environmental Microbiology 62:3662 - 366 PubMed
  • de Ruyter, P.G., Kuipers, O.P., Beerthuyzen, M.M., Alen - Boerrigter, I. and de Vos, W.M. (1996a). Functional analysis of promoters in the nisin gene cluster of Lactococcus lactis. Journal of Bacteriology 178, 3434-3439 PubMed
  • Gasson, M.J. (1983). Plasmid complements of Streptococcus lactis NCDO 712 and other lactic streptococci after protoplast - induced curing. Journal of Bacteriology 154, 1 - 9 PubMed
  • Lei, H. et al. (2015). Protective immunity against influenza H5N1 virus challenge in chickens by oral administration of recombinant Lactococcus lactis expressing neuraminidase. BMC Veterinary Research (2015) 11:85 PubMed
  • Novotny, R., Scheberle, A., Giry-Laterriere, M., Messner, P., Schäffer, C., (2005). Gene cloning, functional expression and secretion of the S-layer protein SgsE from Geobacillus stearothermophilus NRS 2004/3a in Lactococcus lactis. FEMS Microbiol. Lett., 242, 27 - 35 PubMed
  • Platteeuw, C., I. van Alen-Boerrigter, S. van Schalkwijk, and W. M. de Vos. 1996. Food grade cloning and expression system for Lactococcus lactis. Applied and Environmental Microbiology 62:1008 - 1013 PubMed
  • Platteeuw, C., Simons. G., and W. M. de Vos. 1994. Use of the Escherichia coli beta - glucuronidase (gusA) gene as a reporter for analyzing promoters in lactis acid bacteria. Applied and Environmental Microbiology 60 : 587 - 593 PubMed
  • Ravn, P., Arnau, J., Madsen, S. M., Vrang, A., and Israelsen, H. (Aug. 2003). Optimization of signal peptide SP310 for heterologous protein production in Lactococcus lactis. Microbiology 149: 2193 - 2201 PubMed
  • van Asseldonk, M., Rutten, G., Oteman, M., Siezen, R.J., de Vos, W.M., Simons, G. (Oct. 1990). Cloning of usp45, a gene encoding a secreted protein from Lactococcus lactis subsp. lactis MG1363. Gene. 95 (1): 155 - 60 PubMed
  • Vos, P., Simons, G., Siezen, R.J., and de Vos, W. M. (Aug. 1989). Primary structure and organization of the gene for a procaryotic, cell envelope-located serine proteinase . J. Biol. Chem. 264 13579 - 13585 PubMed
  • Wegkamp, A., van Oorschoot, W., de Vos, W. M., and Smid, E. J. (2007). Characterization of the role of para-aminobenzoic acid biosynthesis in folate production by Lactococcus lactis. Applied and Environmental Microbiology 73:2673 - 2681 PubMed