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pTP1172 vector (V001111)

Price Information

Cat No. Plasmid Name Availability Add to cart
V001111 pTP1172 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:
pTP1172
Antibiotic Resistance:
Kanamycin
Length:
4672 bp
Type:
Bacterial Expression
Replication origin:
ori
Copy Number:
High Copy

pTP1172 vector Vector Map

pTP11724672 bp600120018002400300036004200T5 promoterlac operatorRBSlambda t0 terminatorKanRrrnB T1 terminatorCAP binding sitelacIlacIq promoterpGEX 3'bomL4440oripBRforEco

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.

pTP1172 vector Sequence

Copy Sequence

Download GeneBank File(.gb)

LOCUS       pTP1172.        4672 bp DNA     circular SYN 13-MAY-2021
DEFINITION  Bacterial expression plasmid of anti-mouse kappa chain nanobody 
            TP1170 (3x Cysteines).
ACCESSION   .
VERSION     .
KEYWORDS    pTP1172
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4672)
  AUTHORS   Pleiner T,  Bates M,  Gorlich D
  TITLE     A toolbox of anti–mouse and anti–rabbit IgG secondary nanobodies
  JOURNAL   J Cell Biol Dec 2017, jcb.201709115
REFERENCE   2  (bases 1 to 4672)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 4672)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Cell Biol
            Dec 2017, jcb.201709115"
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..4672
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..45
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA
                     polymerase, with embedded lac operator"
     protein_bind    53..69
                     /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)."
     RBS             88..99
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     terminator      908..1002
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             1061..1873
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
     terminator      2095..2181
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     protein_bind    complement(2240..2261)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             complement(2277..3356)
                     /label=lacI
                     /note="lac repressor"
     promoter        complement(3357..3434)
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the
                     promoter boosts expression of the lacI gene about 10-fold."
     primer_bind     complement(3518..3540)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3626..3766
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3781..3798)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3952..4540)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4631..4649
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward
                     primer"