Last data update: 05 March 2021 04:23 CET
Plasmid name: pKNG101 (LMBP 5246)
|Price category:||Cat. 1 (cf. price list)|
|Status:||GeneCorner core plasmid|
(View with Genome Compiler)
|Cloned DNA:||Bacillus subtilis levansucrase cDNA (sacB, GeneID 936413)
Escherichia coli insertion sequence IS1 insB
|Selection marker:||Streptomycin (Sm)|
|Replicon:||Escherichia coli plasmid R6K origin; defective pir gene
Broad-host-range plasmid RK2/RP4 origin of transfer (oriT)
|Host range:||Escherichia coli; use strains supplying the R6K pir function
Gram-negative bacterial strains
|Parental clone:||pJRD215; pKNG100|
|Further information:||pKNG101 is a mobilizable suicide vector that facilitates the positive selection of double recombination events in Gram-negative bacteria.
The plasmid contains: (i) the defective pir(-) origin of replication of the plasmid R6K, (ii) the RK2/RP4 origin of transfer, (iii) the strA and strB genes of RSF1010 encoding streptomycin phosphotransferase as a selection marker for integration, (iv) the sucrose-inducible sacB gene of B. subtilis as a positive selection marker for excision and (v) a multiple cloning site.
The plasmid contains two XbaI sites; the one located at the 3' end of the streptomycin resistance gene only cuts in dam-negative strains.
Take a low-copy-plasmid yield into account while choosing a plasmid DNA isolation procedure.
Introduce the plasmid by electroporation, or through conjugation, using plasmid RK2/RP4 transfer functions (provided in trans by RK2/RP4, RK2/RP4-derived vectors, other replicons carrying RK2/RP4 transfer functions (e.g. pRK2073) or strains expressing the tra genes of RK2/RP4).
The nucleotide sequence of this plasmid corresponds with the EMBL Nucleotide Sequence Database accession number LT726981.1.
The nucleotide sequence was compiled based on the publications mentioned in 'Plasmid reference' and 'Related reference'. The fragment containing the R6K origin and the RK2/RP4 replicon was deduced from the pMRS101 sequencing results obtained at GeneCorner.
|EMBL Accession number:||LT726981.1, view at EMBL, GenBank, DDBJ|
|Latest sequence update:||02/05/2000|
|Authenticity test:||Restriction enzyme pattern analysed at GeneCorner: BamHI, BglI, EcoRI, KpnI, KpnI/NcoI, NcoI, NsiI, NsiI/BglI, NsiI/NcoI, SacI/SmaI, SmaI, SmaI/XmnI and XmnI.|
|History of deposit:||This plasmid was deposited by Prof. Dr G. Cornelis(1).
(1) Microbial Pathogenesis Unit, UCL-ICP, Brussels, Belgium
|Plasmid reference:||Kaniga et al., Gene 109 (1991), 137-141 [PMID: 1756974]
|Related plasmid reference:||Kaniga et al., Gene 100 (1991), 201-205 [PMID: 1647354]
Lathe et al., Gene 57 (1987), 193-201 [PMID: 2826295]
Steinmetz et al., Mol. Gen. Genet. 200 (1985), 220-228 [PMID: 2993818]
Poelwijk et al., Cell 146 (2011), 462-470 [PMID: 21802129] [DOI: 10.1016/j.cell.2011.06.035]
Qiu et al., FEMS Microbiol. Lett. 363 (2016), fnw035 [PMID: 26884480] [DOI: 10.1093/femsle/fnw035]
Ruisheng et al. 27 (2011), 727-730 [DOI: 10.1007/s11274-010-0498-0]
Martinez-Garcia et al., Arch. Microbiol. 175 (2001), 395-404 [PMID: 11491080]
|Restricted distribution:||- BCCM MTA|
|Distributed as:||H/P active culture or plasmid DNA|
|Host for distribution:||Escherichia coli K12 CC118(λpir)|
|Host reference:||Herrero et al., J. Bacteriol. 172 (1990), 6557-6567 [PMID: 2172216]
|Cultivation medium:||LB-Lennox + streptomycin (25 μg/ml)|
|Cultivation remark:||Take a low-copy-plasmid yield into account while choosing a plasmid DNA isolation procedure.|
|Other culture collection numbers:||NCCB 3314
Note: Up-to-date, validated data are enclosed with the biological material. Nevertheless, these data are a snapshot at a given moment; further updates are always possible.