Price Information
| Cat No. | Plasmid Name | Availability | Buy one, get one free! (?) |
|---|---|---|---|
| V012781 | pBAD43 | In stock, instant shipping |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
pBAD43 serves as a recombinant vector backbone, providing arabinose-inducible pBAD promoter regulation of VagD expression. It incorporates a spectinomycin resistance gene for auxiliary screening, whilst its empty vector can also function as a negative control to exclude interference.
- Vector Name:
- pBAD43
- Antibiotic Resistance:
- Spectinomycin
- Length:
- 6187 bp
- Type:
- E.coli expression plasmid
- Replication origin:
- pSC101 ori
- Source/Author:
- Beckwith Lab
- Copy Number:
- Low Copy
- Promoter:
- araBAD
- Cloning Method:
- Enzyme Cut
- 5' Primer:
- pBAD-F: ATGCCATAGCATTTTTATCC
- 3' Primer:
- pBAD-R: gatttaatctgtatcagg
- Growth Strain(s):
- DH10B
- Growth Temperature:
- 30℃
- Expression Method:
- L-arabinose Induced
pBAD43 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
- Duprilot M, Decre D, Genel N, Drieux L, Sougakoff W, Arlet G. Diversity and functionality of plasmid-borne VagCD toxin-antitoxin systems of Klebsiella pneumoniae. J Antimicrob Chemother. 2017 May 1;72(5):1320-1326. doi: 10.1093/jac/dkw569. PMID: 28119479.
pBAD43 vector Sequence
LOCUS Exported 6187 bp DNA circular SYN 06-DEC-2025
DEFINITION synthetic circular DNA.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6187)
TITLE Direct Submission
REFERENCE 2 (bases 1 to 6187)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 6187)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"
COMMENT SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..6187
/mol_type="other DNA"
/organism="synthetic DNA construct"
CDS complement(194..1069)
/codon_start=1
/label=araC
/note="L-arabinose regulatory protein"
/translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
EKVNDVAVKLS"
promoter 1096..1380
/label=araBAD promoter
/note="promoter of the L-arabinose operon of E. coli; the
araC regulatory gene is transcribed in the opposite
direction (Guzman et al., 1995)"
terminator 1664..1750
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
terminator 1842..1869
/label=rrnB T2 terminator
/note="transcription terminator T2 from the E. coli rrnB
gene"
primer_bind complement(1953..1969)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
CDS complement(2495..3283)
/codon_start=1
/label=SmR
/note="aminoglycoside adenylyltransferase (Murphy, 1985)"
/translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKEVAADWAMERLPAQYQP
VIREARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
CDS complement(4323..5270)
/codon_start=1
/label=Rep101
/note="RepA protein needed for replication with the pSC101
origin"
/translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
rep_origin complement(5318..5540)
/direction=LEFT
/label=pSC101 ori
/note="low-copy replication origin that requires the Rep101
protein"
protein_bind 6107..6128
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 6143..6173
/label=lac promoter
/note="promoter for the E. coli lac operon"