Basic Vector Information
- Vector Name:
- pOSIP-KC
- Antibiotic Resistance:
- Kanamycin
- Length:
- 6970 bp
- Type:
- Cloning Vectors
- Replication origin:
- R6K γ ori
- Source/Author:
- St-Pierre F, Cui L, Priest DG, Endy D, Dodd IB, Shearwin KE.
- Copy Number:
- High copy number
pOSIP-KC vector Vector Map
Plasmid Resuspension Protocol:
1. Centrifuge at 5,000×g for 5 min.
2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.
3. Close the tube and incubate for 10 minutes at room temperature.
4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.
5.Store the plasmid at -20 ℃.
pOSIP-KC vector Sequence
LOCUS pOSIP-KC. 6970 bp DNA circular SYN 01-JAN-1980
DEFINITION Prokaryotic one-step cloning and chromosomal integration vector
encoding a kanamycin resistance marker and the phage phi-C31
integrase.
ACCESSION .
VERSION .
KEYWORDS pOSIP-KC.
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6970)
AUTHORS St-Pierre F, Cui L, Priest DG, Endy D, Dodd IB, Shearwin KE.
TITLE One-step cloning and chromosomal integration of DNA.
JOURNAL ACS Synth Biol 2013;2:537-41.
PUBMED 24050148
REFERENCE 2 (bases 1 to 6970)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 6970)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "ACS Synth
Biol"; date: "2013"; volume: "2"; pages: "537-41"
COMMENT SGRef: number: 2; type: "Journal Article"
COMMENT Because of the toxic ccdB gene, this vector must be propagated in a
permissive strain such as DB3.1(TM).
FEATURES Location/Qualifiers
source 1..6970
/mol_type="other DNA"
/organism="synthetic DNA construct"
misc_feature 1..75
/label=MCS 1
/note="MCS 1"
/note="multiple cloning site, part 1"
CDS 142..444
/label=ccdB
/note="CcdB, a bacterial toxin that poisons DNA gyrase"
rep_origin 477..1065
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
misc_feature 1125..1145
/label=BioBrick suffix
/note="universal suffix for all parts"
terminator 1200..1227
/label=rrnB T2 terminator
/note="transcription terminator T2 from the E. coli rrnB
gene"
terminator complement(1276..1307)
/label=tonB terminator
/note="bidirectional E. coli tonB-P14 transcription
terminator"
protein_bind 1384..1417
/label=FRT (minimal)
/note="supports FLP-mediated excision but not integration
(Turan and Bode, 2011)"
CDS complement(1427..2137)
/label=lambda repressor (ts)
/note="temperature-sensitive variant of the phage lambda
repressor"
CDS 2272..4086
/label=phage phi-C31 integrase
/note="integrase from phage phi-C31"
CDS 4224..5036
/label=KanR
/note="aminoglycoside phosphotransferase"
rep_origin 5161..5549
/label=R6K gamma ori
/note="gamma replication origin from E. coli plasmid R6K;
requires the R6K initiator protein pi for replication"
protein_bind 5560..5607
/label=FLP recombinase from the Saccharomyces
cerevisi
/bound_moiety="FLP recombinase from the Saccharomyces
cerevisiae 2u plasmid"
/note="FRT"
/note="FLP-mediated recombination occurs in the 8-bp core
sequence TCTAGAAA (Turan and Bode, 2011)."
protein_bind complement(5676..5775)
/label=phage phi-C31 attP
/note="attachment site of phage phi-C31"
terminator complement(5860..6106)
/label=lambda tL3 terminator
/note="transcription terminator tL3 from phage lambda"
terminator 6428..6514
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
terminator 6611..6697
/gene="Escherichia coli rrnB"
/label=Escherichia coli rrnB terminator
/note="rrnB T1 terminator"
/note="transcription terminator T1 from the E. coli rrnB
gene"
terminator complement(6797..6840)
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
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