pCDH-CAG-MCS-EF1-Puro vector (Cat. No.: V013654)
Note: pCDH-CAG-MCS-EF1-Puro is a lentiviral expression vector. It utilizes the CAG promoter for high-level gene expression in mammalian cells and contains a puromycin resistance marker (Puro) for selection. The vector is designed for generating stable cell lines and is maintained in bacteria with ampicillin resistance.
- Name:
- pCDH-CAG-MCS-EF1-Puro
- Antibiotic Resistance:
- Ampicillin
- Length:
- 8773 bp
- Type:
- Protein expression
- Replication origin:
- ori
- Host:
- Mammalian cells, Lentivirus
- Selection Marker:
- Puro
- Promoter:
- CAG
- Growth Strain(s):
- Stbl3
- Growth Temperature:
- 37℃
Resources
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.
pCDH-CAG-MCS-EF1-Puro vector (Cat. No.: V013654) Sequence
LOCUS Exported 8773 bp DNA circular SYN 30-DEC-2025
DEFINITION Exported.
ACCESSION V013654
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 8773)
TITLE Direct Submission
REFERENCE 2 (bases 1 to 8773)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..8773
/mol_type="other DNA"
/organism="synthetic DNA construct"
source 5981..6001
/mol_type="other DNA"
/organism="synthetic DNA construct"
promoter 6..232
/label=RSV promoter
/note="Rous sarcoma virus enhancer/promoter"
LTR 233..413
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 460..585
/label=HIV-1 Psi
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 1078..1311
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 1496..1540
/label=gp41 peptide
/note="antigenic peptide corresponding to amino acids 655
to 669 of the HIV envelope protein gp41 (Lutje Hulsik et
al., 2013)"
CDS 1689..1730
/label=Protein Tat
/note="Protein Tat from Human immunodeficiency virus type 1
group M subtype B (isolate WMJ22). Accession#: P12509"
misc_feature 1807..1923
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
intron 2605..3621
/label=chimeric intron
/note="chimera between introns from chicken beta-actin and
rabbit beta-globin"
promoter 3733..3944
/label=EF-1-alpha core promoter
/note="core promoter for human elongation factor
EF-1-alpha"
LTR 3957..4225
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from human
T-cell leukemia virus (HTLV) type 1"
CDS 4252..4848
/label=PuroR
/note="puromycin N-acetyltransferase"
misc_feature 4852..5440
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
LTR 5514..5747
/label=3' LTR (Delta-U3)
/note="self-inactivating 3' long terminal repeat (LTR) from
HIV-1"
polyA_signal 5819..5953
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
rep_origin 5959..6115
/label=SV40 ori
/note="SV40 origin of replication"
rep_origin 5981..6001
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind complement(6149..6164)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind complement(6172..6188)
/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(6196..6226)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind complement(6241..6262)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
rep_origin complement(6550..7138)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(7312..8169)
/label=AmpR
/note="beta-lactamase"
promoter complement(8170..8274)
/label=AmpR promoter
primer_bind 8748..8764
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"