Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V006300 | pFAP1 | In stock, 1 week for quality controls |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
Apramycin resistance is obtained after FLP-FRT Recombination.
- Vector Name:
- pFAP1
- Antibiotic Resistance:
- Ampicillin
- Length:
- 3905 bp
- Type:
- Apramycin resistance FRT vector
- Replication origin:
- ori
- Source/Author:
- Choi KH, Gaynor JB, White KG, Lopez C, Bosio CM, Karkhoff-Schweizer RR, Schweizer HP.
pFAP1 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.
pFAP1 vector Sequence
LOCUS 40924_19691 3905 bp DNA circular SYN 18-DEC-2018
DEFINITION Apramycin resistance FRT vector pFAP1, complete sequence.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 3905)
AUTHORS Choi KH, Gaynor JB, White KG, Lopez C, Bosio CM, Karkhoff-Schweizer
RR, Schweizer HP.
TITLE A Tn7-based broad-range bacterial cloning and expression system
JOURNAL Nat. Methods 2 (6), 443-448 (2005)
PUBMED 15908923
REFERENCE 2 (bases 1 to 3905)
AUTHORS Choi K-H., Gaynor JB, White KG, Lopez C, Bosio CM,
Karkhoff-Schweizer RR, Schweizer HP.
TITLE Direct Submission
JOURNAL Submitted (12-JUL-2005) Microbiology, Immunology and Pathology,
Colorado State University, Fort Collins, CO 80523, USA
REFERENCE 3 (bases 1 to 3905)
TITLE Direct Submission
REFERENCE 4 (bases 1 to 3905)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Nat.
Methods"; date: "2005"; volume: "2"; issue: "6"; pages: "443-448"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
(12-JUL-2005) Microbiology, Immunology and Pathology, Colorado State
University, Fort Collins, CO 80523, USA"
COMMENT SGRef: number: 3; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..3905
/mol_type="other DNA"
/organism="synthetic DNA construct"
primer_bind 379..395
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
misc_feature complement(399..455)
/label=MCS
/note="pUC18/19 multiple cloning site"
protein_bind 464..511
/label=FRT
/note="FLP-mediated recombination occurs in the 8-bp core
sequence TCTAGAAA (Turan and Bode, 2011)."
CDS complement(613..1413)
/codon_start=1
/label=ApmR
/note="aminoglycoside 3-N-acetyltransferase type IV"
/translation="MSSAVECNVVQYEWRKAELIGQLLNLGVTPGGVLLVHSSFRSVRP
LEDGPLGLIEALRAALGPGGTLVMPSWSGLDDEPFDPATSPVTPDLGVVSDTFWRLPNV
KRSAHPFAFAAAGPQAEQIISDPLPLPPHSPASPVARVHELDGQVLLLGVGHDANTTLH
LAELMAKVPYGVPRHCTILQDGKLVRVDYLENDHCCERFALADRWLKEKSLQKEGPVGH
AFARLIRSRDIVATALGQLGRDPLIFLHPPEAGCEECDAARQSIG"
misc_feature 1551..1598
/label=FRT, Flp recombinase target site
/note="FRT, Flp recombinase target site"
protein_bind 1551..1598
/label=FRT
/bound_moiety="FLP recombinase from the Saccharomyces
cerevisiae 2u plasmid"
/note="FLP-mediated recombination occurs in the 8-bp core
sequence TCTAGAAA (Turan and Bode, 2011)."
misc_feature 1615..1671
/label=MCS
/note="pUC18/19 multiple cloning site"
primer_bind complement(1684..1700)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind complement(1708..1724)
/label=lac operator
/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)."
promoter complement(1732..1762)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind complement(1777..1798)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
rep_origin complement(2086..2674)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(2848..3705)
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
promoter complement(3706..3810)
/label=AmpR promoter