Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V000381 | pJKR-L-tetR | In stock, 1 week for quality controls |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
- Vector Name:
- pJKR-L-tetR
- Antibiotic Resistance:
- Ampicillin
- Length:
- 5093 bp
- Type:
- Bacterial Expression
- Replication origin:
- pSC101 ori
- Copy Number:
- Low Copy
- Promoter:
- PLtetO-1
pJKR-L-tetR 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.
pJKR-L-tetR vector Sequence
LOCUS 40924_26181 5093 bp DNA circular SYN 13-MAY-2021
DEFINITION Tetracycline biosensor. Low copy plasmid that expresses GFP
proportional to tetracycline concentration..
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5093)
AUTHORS Rogers JK, Guzman CD, Taylor ND, Raman S, Anderson K, Church GM
TITLE Synthetic biosensors for precise gene control and real-time
monitoring of metabolites.
JOURNAL Nucleic Acids Res. 2015 Jul 7. pii: gkv616.
PUBMED 26152303
REFERENCE 2 (bases 1 to 5093)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 5093)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Nucleic
Acids Res. 2015 Jul 7. pii: gkv616."
COMMENT SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..5093
/mol_type="other DNA"
/organism="synthetic DNA construct"
CDS 59..916
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
CDS complement(1076..2023)
/codon_start=1
/label=Rep101
/note="RepA protein needed for replication with the pSC101
origin"
/translation="MSKLVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
LSDMQSKYDLNGSFSWLTQKQRTTLENILAKHGRI"
rep_origin complement(2071..2293)
/direction=LEFT
/label=pSC101 ori
/note="low-copy replication origin that requires the Rep101
protein"
primer_bind complement(2785..2802)
/label=pBAD Reverse
/note="For vectors with E. coli araBAD promoter, reverse
primer"
terminator 2955..3041
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
promoter 3203..3275
/label=PLtetO-1 promoter
/note="modified phage lambda PL promoter with tet operator
sites (Lutz and Bujard, 1997)"
CDS 3313..4026
/codon_start=1
/label=superfolder GFP
/note="GFP variant that folds robustly even when fused to
poorly folded proteins (Pedelacq et al., 2006)"
/translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
FVTAAGITHGMDELYK"
protein_bind complement(4055..4076)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
RBS 4319..4327
/label=Shine-Dalgarno sequence
/note="full consensus sequence for ribosome-binding sites
upstream of start codons in E. coli; complementary to a
region in the 3' end of the 16S rRNA (Chen et al., 1994)"
CDS 4335..4955
/codon_start=1
/label=TetR
/note="tetracycline repressor TetR"
/translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
terminator 5012..5059
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"