Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V017067 | pAAVS1P-iCAG.copGFP | 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
- Vector Name:
- pAAVS1P-iCAG.copGFP
- Antibiotic Resistance:
- Kanamycin
- Length:
- 10760 bp
- Type:
- Gene knockin
- Replication origin:
- ori
- Host:
- Mammalian cells
- Selection Marker:
- Puro
- Promoter:
- CAG
- Growth Strain(s):
- Stbl3
- Growth Temperature:
- 37℃
pAAVS1P-iCAG.copGFP vector 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.
pAAVS1P-iCAG.copGFP vector Sequence
LOCUS 62056_2620 10760 bp DNA circular SYN 01-JAN-1980
DEFINITION synthetic circular DNA.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 10760)
AUTHORS .
TITLE Direct Submission
FEATURES Location/Qualifiers
source 1..10760
/mol_type="other DNA"
/organism="synthetic DNA construct"
CDS 114..920
/codon_start=1
/label=KanR
/note="aminoglycoside phosphotransferase"
/translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
rep_origin 1101..1689
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
protein_bind 1977..1998
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 2013..2043
/label=lac promoter
/note="promoter for the E. coli lac operon"
misc_feature 2112..2915
/label=HA-L
/note="left homology arm from the adeno-associated virus
integration site (AAVS1) within intron 1 of the human
PPP1R12C gene"
misc_feature 2958..2983
/label=SA
/note="splice acceptor site"
CDS 3007..3060
/codon_start=1
/label=T2A
/note="2A peptide from Thosea asigna virus capsid protein"
/translation="EGRGSLLTCGDVEENPGP"
CDS 3064..3660
/codon_start=1
/label=PuroR
/note="puromycin N-acetyltransferase"
/translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
polyA_signal 3664..3888
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
protein_bind complement(3889..3922)
/label=loxP
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (ATGTATGC) (Shaw et al., 2021)."
protein_bind complement(5166..5199)
/label=lox2272
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are
compatible with each other, but incompatible with loxP or
loxN sites (Lee and Saito, 1988)."
intron 5854..6871
/label=chimeric intron
/note="chimera between introns from chicken beta-actin and
rabbit beta-globin"
CDS 6956..7621
/codon_start=1
/label=CopGFP
/note="green fluorescent protein 2 from Pontellina plumata,
also known as ppluGFP2 (Shagin et al., 2004)"
/translation="LPAMEIECRITGTLNGVEFELVGGGEGTPKQGRMTNKMKSTKGAL
TFSPYLLSHVMGYGFYHFGTYPSGYENPFLHAINNGGYTNTRIEKYEDGGVLHVSFSYR
YEAGRVIGDFKVVGTGFPEDSVIFTDKIIRSNATVEHLHPMGDNVLVGSFARTFSLRDG
GYYSFVVDSHMHFKSAIHPSILQNGGPMFAFRRVEELHSNTELGIVEYQHAFKTPIAFA
"
polyA_signal 7848..7903
/label=beta-globin poly(A) signal
/note="rabbit beta-globin polyadenylation signal (Gil and
Proudfoot, 1987)"
protein_bind 8236..8269
/label=loxP511
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (ATGTATAC) (Shaw et al., 2021). loxP511 sites are
compatible with each other, but incompatible with wild-type
loxP sites."