Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V002803 | ptCC9 | 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
- Vector Name:
- ptCC9
- Antibiotic Resistance:
- Ampicillin
- Length:
- 8857 bp
- Type:
- Shuttle vector
- Replication origin:
- ori
- Source/Author:
- Stukenberg D, Zauner S, Dell'Aquila G, Maier UG.
- Growth Strain(s):
- Top10
- Growth Temperature:
- 37℃
ptCC9 vector Map
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.
ptCC9 vector Sequence
LOCUS 40924_42714 8857 bp DNA circular SYN 18-DEC-2018
DEFINITION Shuttle vector ptCC9, complete sequence.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 8857)
AUTHORS Stukenberg D, Zauner S, Dell'Aquila G, Maier UG.
TITLE Optimizing CRISPR/Cas9 for the Diatom Phaeodactylum tricornutum
JOURNAL Front Plant Sci 9, 740 (2018)
PUBMED 29928285
REFERENCE 2 (bases 1 to 8857)
AUTHORS Stuckenberg D, Zauner S.
TITLE Direct Submission
JOURNAL Submitted (29-MAR-2018) Philipps-University Marburg, Cellbiology,
Karl-von-Frisch Str. 8, Marburg 35043, Deutschland
REFERENCE 3 (bases 1 to 8857)
TITLE Direct Submission
REFERENCE 4 (bases 1 to 8857)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Front Plant
Sci 9, 740 (2018)"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
(29-MAR-2018) Philipps-University Marburg, Cellbiology,
Karl-von-Frisch Str. 8, Marburg 35043, Deutschland"
COMMENT SGRef: number: 3; type: "Journal Article"
COMMENT ##Assembly-Data-START##
Sequencing Technology :: Sanger dideoxy sequencing
##Assembly-Data-END##
FEATURES Location/Qualifiers
source 1..8857
/mol_type="other DNA"
/organism="synthetic DNA construct"
regulatory 1..415
/note="nitrate reductase; derived from Phaeodactylum
tricornutum"
/regulatory_class="promoter"
CDS 431..496
/codon_start=1
/product="three tandem FLAG(R) epitope tags, followed by an
enterokinase cleavage site"
/label=3xFLAG
/translation="DYKDHDGDYKDHDIDYKDDDDK"
CDS 503..523
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 548..4648
/label=Cas9
/note="Cas9 (Csn1) endonuclease from the Streptococcus
pyogenes Type II CRISPR/Cas system"
CDS 4649..4696
/codon_start=1
/product="bipartite nuclear localization signal from
nucleoplasmin"
/label=nucleoplasmin NLS
/translation="KRPAATKKAGQAKKKK"
regulatory 4700..4972
/note="nitrate reductase; derived from Phaeodactylum
tricornutum"
/regulatory_class="terminator"
regulatory 5010..5288
/note="U6-RNA promoter; derived from Phaeodactylum
tricornutum"
/regulatory_class="promoter"
misc_RNA 5310..5385
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
regulatory 5386..5723
/note="U6-RNA terminator; derived from Phaeodactylum
tricornutum"
/regulatory_class="terminator"
regulatory 5724..5916
/note="fcpB promoter; derived from Phaeodactylum
tricornutum"
/regulatory_class="promoter"
CDS 5917..6288
/label=BleoR
/note="antibiotic-binding protein"
regulatory 6292..6532
/note="fcpB teminator; derived from Phaeodactylum
tricornutum"
/regulatory_class="terminator"
promoter complement(6543..6561)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
rep_origin complement(6819..7407)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(7581..8438)
/label=AmpR
/note="beta-lactamase"
promoter complement(8439..8543)
/label=AmpR promoter