pSET2 vector (V003385)

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Price Information

Cat No.	Plasmid Name	Availability	Add to cart
V003385	pSET2	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

pSET1, pSET2, and pSET3 carry cat, spc, and both of these genes, respectively, as selectable markers. These vectors could be introduced into S. suis, E. coli, Salmonella typhimurium, S. pneumoniae, and S. equi ssp. equi by electrotransformation.

Vector Name:: pSET2

Antibiotic Resistance:: Spectinomycin

Length:: 5016 bp

Type:: Shuttle vector

Source/Author:: Takamatsu D, Osaki M, Sekizaki T.

Promoter:: lac

Growth Temperature:: 37℃

pSET2 vector Map

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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

Zheng C, Wei M, Qiu J, Li J. A Markerless Gene Deletion System in Streptococcus suis by Using the Copper-Inducible Vibrio parahaemolyticus YoeB Toxin as a Counterselectable Marker. Microorganisms. 2021 May 19;9(5):1095.

pSET2 vector Sequence

LOCUS       Exported                5016 bp DNA     circular SYN 12-MAR-2024
DEFINITION  Shuttle vector pSET2 DNA, complete sequence.
ACCESSION   AB042430
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5016)
  AUTHORS   Takamatsu D, Osaki M, Sekizaki T.
  TITLE     Construction and characterization of Streptococcus suis-Escherichia 
            coli shuttle cloning vectors
  JOURNAL   Plasmid 45 (2), 101-113 (2001)
  PUBMED    11322824
REFERENCE   2  (bases 1 to 5016)
  AUTHORS   Takamatsu D, Osaki M, Sekizaki T.
  TITLE     Direct Submission
  JOURNAL   Submitted (08-MAY-2000) Daisuke Takamatsu, National Institute of 
            Animal Health, Laboratory of Molecular Bacteriology; Kannondai 
            3-1-1, Tukuba, Ibaraki 305-0856, Japan 
            (E-mail:p1013dt@niah.affrc.go.jp, Tel:81-298-38-7743, 
            Fax:81-298-38-7743)
REFERENCE   3  (bases 1 to 5016)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Plasmid"; 
            date: "2001"; volume: "45"; issue: "2"; pages: "101-113"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
            (08-MAY-2000) Daisuke Takamatsu, National Institute of Animal
            Health, Laboratory of Molecular Bacteriology"; pages: 
            "81-298-38-7743"
FEATURES             Location/Qualifiers
     source          1..5016
                     /lab_host="Streptococcus suis, Escherichia coli"
                     /mol_type="other DNA"
                     /label=synonym:Shuttle Cloning Vector pSET2
                     /note="synonym:Shuttle Cloning Vector pSET2"
                     /db_xref="taxon:125568"
                     /organism="Shuttle vector pSET2"
     gene            156..176
                     /gene="dso"
                     /label=dso
     misc_feature    156..176
                     /gene="dso"
                     /label=double-stranded replication origin
                     /note="double-stranded replication origin"
     gene            367..504
                     /gene="copG"
                     /label=copG
     CDS             367..504
                     /codon_start=1
                     /gene="copG"
                     /product="CopG, transcriptional repressor protein"
                     /function="transcriptional repressor protein"
                     /label=copG
                     /protein_id="BAB03239.1"
                     /translation="MKKRLTITLSESVLENLEKMAKEMGLSKSALISVALENYKKGQVK
                     "
     gene            566..1246
                     /gene="repB"
                     /label=repB
     CDS             566..1246
                     /codon_start=1
                     /gene="repB"
                     /product="RepB, replication initiation¨Ctermination
                     protein."
                     /function="The plasmid-encoded initiator of replication.The
                     protein introduces a strand- and site-specific nick in the 
                     double-stranded replication origin, on supercoiled DNA."
                     /label=repB
                     /protein_id="BAB03240.1"
                     /translation="MAKEKARYFTFLLYPESIPSDWELKLELLGVPIAVSPLHDRDKSD
                     VEGQQYKKPHYHVIYVSKNPVTADSVRMKIKRSLGDNSVALVQIIRTSIENTYLYLTHE
                     SKDAIEKKKHVYDKADITLLSNFDIDRYITLDVEEKDDMLNEVCDLIDEYDIANMRELR
                     RFIKLHGAEHGLPSIKIINSVLRSHTGLIRLYFDAVYQERKYGSNIPNVNQETGEILSE
                     DSDD"
     gene            complement(1703..2470)
                     /gene="add9"
                     /label=add9
     CDS             complement(1703..2470)
                     /codon_start=1
                     /gene="add9"
                     /product="spectinomycin adenyltransferase, aso known as
                     aad9."
                     /function="spectinomycin resistance"
                     /label=spc
                     /protein_id="BAB03241.1"
                     /translation="MRRIYLNTYEQINKVKKILRKHLKNNLIGTYMFGSGVESGLKPNS
                     DLDFLVVVSEPLTDQSKEILIQKIRPISKKIGDKSNLRYIELTIIIQQEMVPWNHPPKQ
                     EFIYGEWLQELYEQGYIPQKELNSDLTIMLYQAKRKNKRIYGNYDLEELLPDIPFSDVR
                     RAIMDSSEELIDNYQDDETNSILTLCRMILTMDTGKIIPKDIAGNAVAESSPLEHRERI
                     LLAVRSYLGENIEWTNENVNLTINYLNNRLKKL"
     gene            complement(2872..3195)
                     /gene="lacZ'"
                     /label=lacZ'
     CDS             complement(2872..3195)
                     /codon_start=1
                     /gene="lacZ'"
                     /product="beta-galactosidase alpha fragment"
                     /label=lacZ'
                     /protein_id="BAB03242.1"
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     primer_bind     3105..3121
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    3122..3178
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(3191..3207)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3215..3231
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /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(3239..3269)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3284..3305
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     gene            4111..4252
                     /gene="ssoA"
                     /label=ssoA
     misc_feature    4111..4252
                     /gene="ssoA"
                     /label=single-stranded replication origin
                     /note="single-stranded replication origin"