Price Information
| Cat No. | Plasmid Name | Availability | Buy one, get one free! (?) |
|---|---|---|---|
| V012110 | pSIM6 | In stock, instant shipping |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
Plasmid pSim6 was constructed by recombining the defective prophage into the pSC101 plasmid backbone. A 6,719-bp plasmid characterized by ONT sequencing, which is a low-copy homologous recombination vector. Its replication requires the temperature-sensitive Rep101 protein, resulting in plasmid loss at 37°C, which enables controlled gene expression. Featuring multiple cloning sites and an ampicillin resistance marker, it supports diverse DNA insertions in cell lines like 293/CV-1. As a constitutive vector, it ensures stable, continuous protein production, making it ideal for temperature-regulated gene editing and expression studies.
- Vector Name:
- pSIM6
- Antibiotic Resistance:
- Ampicillin
- Length:
- 6719 bp
- Type:
- Homologous recombinant vectors
- Replication origin:
- pSC101 ori
- Copy Number:
- Low Copy
- Growth Strain(s):
- DH10B
- Growth Temperature:
- 30℃
pSIM6 vector Map
Plasmid 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 ℃.
6. The concentration of plasmid re-measurement sometimes differs from the nominal value, which may be due to the position of the lyophilized plasmid in the tube, the efficiency of the re-dissolution, the measurement bias, and adsorption on the wall of the tube, therefore, it is recommended to transform and extract the plasmid before using it
General Plasmid Transform Protocol
1. Take one 100μl of the competent cells and thaw it on ice for 10min, add 2μl of plasmid, then ice bath for 30min, then heat-shock it at 42℃ for 60s, do not stir, and then ice bath for 2min.
2. Add 900μl of LB liquid medium without antibiotics, and incubate at 37℃ for 45min (30℃ for 1-1.5 hours) with 180rpm shaking.
3. Centrifuge at 6000rpm for 5min, leave only 100μl of supernatant to resuspend the bacterial precipitate and spread it onto the target plasmid-resistant LB plate.
4. Invert the plate and incubate at 37℃ for 14h, or at 30℃ for 20h.
5. Pick a single colony into LB liquid medium, add the corresponding antibiotics, incubate at 220rpm for 14h, and extract the plasmid according to the experimental needs and the instructions of the plasmid extraction kit.
References
- Chan W, Costantino N, Li R, Lee SC, Su Q, Melvin D, Court DL, Liu P. A recombineering based approach for high-throughput conditional knockout targeting vector construction. Nucleic Acids Res. 2007;35(8):e64. doi: 10.1093/nar/gkm163. Epub 2007 Apr 10. PMID: 17426124; PMCID: PMC1885671.
pSIM6 vector Sequence
LOCUS Exported 6719 bp DNA circular SYN 20-JUL-2025
DEFINITION synthetic circular DNA.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6719)
TITLE Direct Submission
REFERENCE 2 (bases 1 to 6719)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 6719)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"
COMMENT SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..6719
/mol_type="other DNA"
/organism="synthetic DNA construct"
source 4578..4580
/mol_type="other DNA"
/organism="synthetic DNA construct"
misc_feature 4..22
/label=Tn5 ME
/note="hyperactive mosaic end for Tn5 transposase
recognition (Reznikoff et al., 2004)"
CDS complement(142..819)
/codon_start=1
/label=Exo
/note="5' to 3' double-stranded DNA exonuclease in the
lambda
Red system"
/translation="MTPDIILQRTGIDVRAVEQGEDAWHKLRLGVITASEVHNVIAKPR
SGKKWPDMKMSYFHTLLAEVCTGVAPEVNAKALAWGKQYENDARTLFEFTSGVNVTESP
IIYRDESMRTACSPDGLCSDGNGLELKCPFTSRDFMKFRLGGFEAIKSAYMAQVQYSMW
VTRKNAWYFANYDPRMKREGLHYVVIERDEKYMASFDEIVPEFIEKMDEALAEIGFVFG
EQWR"
CDS complement(819..1601)
/codon_start=1
/label=Beta
/note="single-stranded DNA binding recombinase in the
lambda Red system"
/translation="MSTALATLAGKLAERVGMDSVDPQELITTLRQTAFKGDASDAQFI
ALLIVANQYGLNPWTKEIYAFPDKQNGIVPVVGVDGWSRIINENQQFDGMDFEQDNESC
TCRIYRKDRNHPICVTEWMDECRREPFKTREGREITGPWQSHPKRMLRHKAMIQCARLA
FGFAGIYDKDEAERIVENTAYTAERQPERDITPVNDETMQEINTLLIALDKTWDDDLLP
LCSQIFRRDIRASSELTQAEAVKALGFLKQKAAEQKVAA"
CDS complement(2615..3472)
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
promoter complement(3473..3544)
/label=AmpR promoter
CDS complement(3660..4370)
/codon_start=1
/label=lambda repressor (ts)
/note="temperature-sensitive variant of the phage lambda
repressor"
/translation="MSTKKKPLTQEQLEDARRLKAIYEKKKNELGLSQESVADKMGMGQ
SGVGALFNGINALNAYNAALLTKILKVSVEEFSPSIAREIYEMYEAVSMQPSLRSEYEY
PVFSHVQAGMFSPELRTFTKGDAERWVSTTKKASDSAFWLEVEGNSMTAPTGSKPSFPD
GMLILVDPEQAVEPGDFCIARLGGDEFTFKKLIRDSGQVFLQPLNPQYPMIPCNESCSV
VGKVIASQWPEETFG"
misc_feature complement(4482..4500)
/label=Tn5 ME
/note="hyperactive mosaic end for Tn5 transposase
recognition (Reznikoff et al., 2004)"
CDS complement(5023..5970)
/codon_start=1
/label=Rep101
/note="RepA protein needed for replication with the pSC101
origin"
/translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
RTVSFTYNQYVQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
rep_origin complement(6018..6240)
/direction=LEFT
/label=pSC101 ori
/note="low-copy replication origin that requires the Rep101
protein"