Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V003722 | pRK442 | In stock, instant shipping |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
- Vector Name:
- pRK442
- Antibiotic Resistance:
- Tetracycline
- Length:
- 9816 bp
- Type:
- Cloning vector
- Replication origin:
- oriV
- Source/Author:
- Scott HN, Laible PD, Hanson DK.
pRK442 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.
pRK442 vector Sequence
LOCUS 40924_37143 9816 bp DNA circular SYN 18-DEC-2018
DEFINITION Cloning vector pRK442, complete sequence.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 9816)
AUTHORS Scott HN, Laible PD, Hanson DK.
TITLE Sequences of versatile broad-host-range vectors of the RK2 family
JOURNAL Plasmid 50 (1), 74-79 (2003)
PUBMED 12826060
REFERENCE 2 (bases 1 to 9816)
AUTHORS Scott HN, Laible PD, Hanson DK.
TITLE Direct Submission
JOURNAL Submitted (18-DEC-2002) Biosciences Division, Argonne National
Laboratory, 9700 South Cass Ave., Argonne, IL 60439, USA
REFERENCE 3 (bases 1 to 9816)
TITLE Direct Submission
REFERENCE 4 (bases 1 to 9816)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Plasmid";
date: "2003"; volume: "50"; issue: "1"; pages: "74-79"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
(18-DEC-2002) Biosciences Division, Argonne National Laboratory,
9700 South Cass Ave., Argonne, IL 60439, USA"
COMMENT SGRef: number: 3; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..9816
/mol_type="other DNA"
/organism="synthetic DNA construct"
rep_origin complement(152..863)
/direction=LEFT
/label=oriV
/note="incP origin of replication"
primer_bind 1373..1389
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(1438..1454)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind complement(1462..1478)
/label=lac operator
/note="The lac repressor binds to the lac operator to
inhibit transcription in E. coli. This inhibition can be
relieved by adding lactose or
isopropyl-beta-D-thiogalactopyranoside (IPTG)."
promoter complement(1486..1516)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind complement(1531..1552)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
CDS complement(1787..2155)
/codon_start=1
/label=traJ
/note="oriT-recognizing protein"
/translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
KQDELGKVMMGVVRPRAEP"
oriT complement(2188..2297)
/direction=LEFT
/label=oriT
/note="incP origin of transfer"
CDS complement(2981..3628)
/codon_start=1
/label=TetR
/note="tetracycline resistance regulatory protein"
/translation="MTKLQPNTVIRAALDLLNEVGVDGLTTRKLAERLGVQQPALYWHF
RNKRALLDALAEAMLAENHTHSVPRADDDWRSFLIGNARSFRQALLAYRDGARIHAGTR
PGAPQMETADAQLRFLCEAGFSAGDAVNALMTISYFTVGAVLEEQAGDSDAGERGGTVE
QAPLSPLLRAAIDAFDEAGPDAAFEQGLAVIVDGLAKRRLVVRNVEGPRKGDD"
CDS 3734..4930
/codon_start=1
/label=TcR
/note="tetracycline efflux protein"
/translation="MKPNIPLIVILSTVALDAVGIGLIMPVLPGLLRDLVHSNDVTAHY
GILLALYALVQFACAPVLGALSDRFGRRPILLVSLAGATVDYAIMATAPFLWVLYIGRI
VAGITGATGAVAGAYIADITDGDERARHFGFMSACFGFGMVAGPVLGGLMGGFSPHAPF
FAAAALNGLNFLTGCFLLPESHKGERRPLRREALNPLASFRWARGMTVVAALMAVFFIM
QLVGQVPAALWVIFGEDRFHWDATTIGISLAAFGILHSLAQAMITGPVAARLGERRALM
LGMIADGTGYILLAFATRGWMAFPIMVLLASGGIGMPALQAMLSRQVDEERQGQLQGSL
AALTSLTSIVGPLLFTAIYAASITTWNGWAWIAGAALYLLCLPALRRGLWSGAGQRADR
"
CDS complement(6209..7354)
/codon_start=1
/label=trfA
/note="trans-acting replication protein that binds to and
activates oriV"
/translation="MNRTFDRKAYRQELIDAGFSAEDAETIASRTVMRAPRETFQSVGS
MVQQATAKIERDSVQLAPPALPAPSAAVERSRRLEQEAAGLAKSMTIDTRGTMTTKKRK
TAGEDLAKQVSEAKQAALLKHTKQQIKEMQLSLFDIAPWPDTMRAMPNDTARSALFTTR
NKKIPREALQNKVIFHVNKDVKITYTGVELRADDDELVWQQVLEYAKRTPIGEPITFTF
YELCQDLGWSINGRYYTKAEECLSRLQATAMGFTSDRVGHLESVSLLHRFRVLDRGKKT
SRCQVLIDEEIVVLFAGDHYTKFIWEKYRKLSPTARRMFDYFSSHREPYPLKLETFRLM
CGSDSTRVKKWREQVGEACEELRGSGLVEHAWVNDDLVHCKR"
CDS complement(8081..9175)
/codon_start=1
/gene="incC"
/product="plasmid maintenance protein"
/label=incC
/protein_id="AAO47421.1"
/translation="MGVIHEETAYRKPVPGGDPGAGSGAADHRDSAGRLSRWEATGDVR
NVAGTDQGRSVASGASRVGRVRGQELARGVRAGNGGSAGTSGVHRPEVGSGRQEKTGNQ
TMKTLVTANQKGGVGKTSTLVHLAFDFFERGLRVAVIDLDPQGNASYTLKDFATGLHAS
KLFGAVPAGGWTETAPAAGDGQAARLALIESNPVLANAERLSLDDARELFGANIKALAN
QGFDVCLIDTAPTLGVGLAAALFAADYVLSPIELEAYSIQGIKKMVTTIANVRQKNAKL
QFLGMVPSKVDARNPRHARHQAELLAAYPKMMIPATVGLRSSIADALASGVPVWKIKKT
AARKASKEVRALADYVFTKMEISQ"
gene complement(8081..9175)
/gene="incC"
/label=incC
CDS complement(8857..9162)
/codon_start=1
/gene="korA"
/product="regulation of gene expression"
/label=korA
/protein_id="AAO47422.1"
/translation="MKKRLTESQFQEAIQGLEVGQQTIEIARGVLVDGKPQATFATSLG
LTRGAVSQAVHRVWAAFEDKNLPEGYARVTAVLPEHQAYIVRKWEADAKKKQETKR"
gene complement(8857..9162)
/gene="korA"
/label=korA