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pDHtsk-BenA vector (V014504)

Price Information

Cat No. Plasmid Name Availability Add to cart
V014504 pDHtsk-BenA In stock, 1 week for quality controls

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Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.

Basic Vector Information

Vector Name:
pDHtsk-BenA
Antibiotic Resistance:
Kanamycin
Length:
10259 bp
Type:
Protein expression
Replication origin:
ori
Host:
Hyphal fungi
Selection Marker:
BenA
Promoter:
NOS
Growth Strain(s):
DH5alpha
Growth Temperature:
37℃

pDHtsk-BenA vector Map

Copy Sequence View Map
pDHtsk-BenA10259 bp50010001500200025003000350040004500500055006000650070007500800085009000950010000T7 promoterT3 promoterNOS promoterTubulin beta chainNOS terminatorT3 promoterlac promoterCAP binding siteRB T-DNA repeatpVS1 StaApVS1 RepApVS1 oriVbomoriKanRLB T-DNA repeatCaMV poly(A) signal

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.

pDHtsk-BenA vector Sequence

LOCUS       V014504                10259 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  Exported.
ACCESSION   V014504
VERSION     V014504
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 10259)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..10259
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        191..209
                     /label="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        294..312
                     /label="T3 promoter"
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        654..837
                     /label="NOS promoter"
                     /note="nopaline synthase promoter"
     CDS             858..2198
                     /gene="tubA"
                     /label="Tubulin beta chain"
                     /note="Tubulin beta chain from Botryotinia fuckeliana.
                     Accession#: P53373"
     terminator      2577..2829
                     /label="NOS terminator"
                     /note="nopaline synthase terminator and poly(A) signal"
     promoter        complement(3175..3193)
                     /label="T3 promoter"
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        complement(3262..3293)
                     /label="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(3308..3329)
                     /label="CAP binding site"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     misc_feature    3697..3721
                     /label="RB T-DNA repeat"
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             5021..5647
                     /label="pVS1 StaA"
                     /note="stability protein from the Pseudomonas plasmid pVS1
                     (Heeb et al., 2000)"
     CDS             6084..7148
                     /label="pVS1 RepA"
                     /note="replication protein from the Pseudomonas plasmid
                     pVS1 (Heeb et al., 2000)"
     rep_origin      7217..7411
                     /label="pVS1 oriV"
                     /note="origin of replication for the Pseudomonas plasmid
                     pVS1 (Heeb et al., 2000)"
     misc_feature    7755..7895
                     /label="bom"
                     /note="basis of mobility region from pBR322"
     rep_origin      complement(8081..8669)
                     /direction=LEFT
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     CDS             complement(8759..9550)
                     /label="KanR"
                     /note="aminoglycoside phosphotransferase"
     misc_feature    9975..9999
                     /label="LB T-DNA repeat"
                     /note="left border repeat from nopaline C58 T-DNA"
     polyA_signal    complement(10077..10251)
                     /label="CaMV poly(A) signal"
                     /note="cauliflower mosaic virus polyadenylation signal"