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pPICZα B vector (V010240)

Price Information

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V010240 pPICZα B In stock, instant shipping

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

Basic Vector Information

pPICZα B vector (V010240) is a Pichia pastoris vector for methanol-inducible expression of a secreted protein. There are other similar expression vectors, such as pPICZα A or pPICZα C For other reading frames.

Vector Name:
pPICZα B
Antibiotic Resistance:
Bleomycin
Length:
3597 bp
Type:
Yeast Plasmids
Replication origin:
ori
Source/Author:
Invitrogen (Life Technologies)
Copy Number:
High copy number
Promoter:
AOX1
Growth Strain(s):
Stbl3
Growth Temperature:
37℃

pPICZα B vector Map

Copy Sequence View Map
pPICZα B3597 bp6001200180024003000AOX1 promoteralpha-factor secretion signalMyc6xHisAOX1 terminatorTEF1 promoterEM7 promoterBleoRCYC1 terminatorori

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

  • Zhao RL, Zhang XM, Jia LN, Song W, Sun YL, Meng XY, Peng XX. pNNS-Conjugated Chitosan Mediated IGF-1 and miR-140 Overexpression in Articular Chondrocytes Improves Cartilage Repair. Biomed Res Int. 2019 Mar 21;2019:2761241.

pPICZα B vector Sequence

LOCUS       pPICZ-B.        3597 bp DNA     circular SYN 13-SEP-2021
DEFINITION  Pichia pastoris vector for methanol-inducible expression of a 
            secreted protein.
ACCESSION   .
VERSION     .
KEYWORDS    pPICZ-B
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3597)
  AUTHORS   Invitrogen (Life Technologies)
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 3597)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 3597)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     SGRef: number: 2; type: "Journal Article"
COMMENT     Linearize with SacI, PmeI, or BstXI for integration at the AOX1 
            locus.
FEATURES             Location/Qualifiers
     source          1..3597
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        2..940
                     /label=AOX1 promoter
                     /note="inducible promoter, regulated by methanol"
     CDS             941..1207
                     /label=alpha-factor secretion signal
                     /note="N-terminal secretion signal from S. cerevisiae 
                     alpha-factor"
     misc_feature    1206..1280
                     /label=MCS
                     /note="MCS"
                     /note="MCS;multiple cloning site"
     CDS             1279..1308
                     /label=Myc
                     /note="Myc (human c-Myc proto-oncogene) epitope tag"
     CDS             1324..1341
                     /label=6xHis
                     /note="6xHis affinity tag"
     terminator      1421..1667
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     promoter        1682..2093
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     promoter        2101..2148
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter"
     CDS             2167..2538
                     /label=BleoR
                     /note="antibiotic-binding protein"
     terminator      2607..2854
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     rep_origin      complement(2929..3517)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"