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pPha-NR vector (V004111)

Price Information

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V004111 pPha-NR In stock, instant shipping

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

Basic Vector Information

Vector Name:
pPha-NR
Antibiotic Resistance:
Ampicillin
Length:
3860 bp
Type:
Shuttle vector
Replication origin:
ori
Source/Author:
Stork S, Moog D, Przyborski JM, Wilhelmi I, Zauner S, Maier UG.
Growth Strain(s):
Top10
Growth Temperature:
37℃

pPha-NR vector Map

Copy Sequence View Map
pPha-NR3860 bp60012001800240030003600nitrate reductase; derived from Phaeodactylum tricornutumMCSnitrate reductase; derived from Phaeodactylum tricornutumfcpB; derived from Phaeodactylum tricornutumBleoRfcpA; derived from Phaeodactylum tricornutumT7 promoteroriAmpRAmpR promoter

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.

pPha-NR vector Sequence

LOCUS       40924_34441        3860 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Shuttle vector pPha-NR, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3860)
  AUTHORS   Stork S, Moog D, Przyborski JM, Wilhelmi I, Zauner S, Maier UG.
  TITLE     Distribution of the SELMA Translocon in Secondary Plastids of Red 
            Algal Origin and Predicted Uncoupling of Ubiquitin-Dependent 
            Translocation from Degradation
  JOURNAL   Eukaryotic Cell 11 (12), 1472-1481 (2012)
  PUBMED    23042132
REFERENCE   2  (bases 1 to 3860)
  AUTHORS   Zauner S, Hempel F.
  TITLE     Inducible shuttle vector
  JOURNAL   Unpublished
REFERENCE   3  (bases 1 to 3860)
  AUTHORS   Zauner S, Hempel F.
  TITLE     Direct Submission
  JOURNAL   Submitted (24-JUN-2011) Cell Biology, Philipps-University Marburg, 
            Karl-von-Frisch Str. 8, Marburg 35032, Germany
REFERENCE   4  (bases 1 to 3860)
  TITLE     Direct Submission
REFERENCE   5  (bases 1 to 3860)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Eukaryotic 
            Cell"; date: "2012"; volume: "11"; issue: "12"; pages: "1472-1481"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 3; type: "Journal Article"; journalName: "Submitted 
            (24-JUN-2011) Cell Biology, Philipps-University Marburg, 
            Karl-von-Frisch Str. 8, Marburg 35032, Germany"
COMMENT     SGRef: number: 4; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..3860
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      1..422
                     /note="nitrate reductase; derived from Phaeodactylum 
                     tricornutum"
                     /regulatory_class="promoter"
     misc_feature    441..497
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     regulatory      498..770
                     /note="nitrate reductase; derived from Phaeodactylum 
                     tricornutum"
                     /regulatory_class="terminator"
     regulatory      771..956
                     /note="fcpB; derived from Phaeodactylum tricornutum"
                     /regulatory_class="promoter"
     CDS             957..1328
                     /label=BleoR
                     /note="antibiotic-binding protein"
     regulatory      1331..1573
                     /note="fcpA; derived from Phaeodactylum tricornutum"
                     /regulatory_class="terminator"
     promoter        complement(1583..1601)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     rep_origin      complement(1859..2447)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(2621..3478)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(3479..3583)
                     /label=AmpR promoter