Basic Vector Information
- Vector Name:
- pLV3-U6-MCS-BsmBI-EcoRI-sgRNA-Cas9-EGFP-Puro
- Antibiotic Resistance:
- Ampicillin
- Length:
- 12993 bp
- Type:
- Genome editing
- Replication origin:
- ori
- Host:
- Mammalian cells, Lentivirus
- Selection Marker:
- Puro
- Promoter:
- U6
- 5' Primer:
- U6-F
- Growth Temperature:
- 37℃
pLV3-U6-MCS-BsmBI-EcoRI-sgRNA-Cas9-EGFP-Puro vector Vector Map
Plasmid Resuspension 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 ℃.
pLV3-U6-MCS-BsmBI-EcoRI-sgRNA-Cas9-EGFP-Puro vector Sequence
LOCUS 62056_15640 12993 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 12993)
AUTHORS .
TITLE Direct Submission
FEATURES Location/Qualifiers
source 1..12993
/mol_type="other DNA"
/organism="synthetic DNA construct"
promoter 1..241
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
misc_RNA 288..363
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
promoter 425..636
/label=EF-1-alpha core promoter
/note="core promoter for human elongation factor
EF-1-alpha"
CDS 661..4764
/label=Cas9
/note="Cas9 (Csn1) endonuclease from the Streptococcus
pyogenes Type II CRISPR/Cas system"
CDS 4813..4836
/label=FLAG
/note="FLAG(R) epitope tag, followed by an enterokinase
cleavage site"
CDS 4903..5619
/label=EGFP
/note="enhanced GFP"
misc_feature 6296..6884
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
LTR 6956..7189
/label=3' LTR (Delta-U3)
/note="self-inactivating 3' long terminal repeat (LTR) from
HIV-1"
polyA_signal 7221..7445
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 7491..7919
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
polyA_signal 8062..8195
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
promoter complement(8280..8310)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind complement(8325..8346)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
rep_origin complement(8634..9222)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(9396..10253)
/label=AmpR
/note="beta-lactamase"
promoter complement(10254..10358)
/label=AmpR promoter
enhancer 10624..11003
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 11004..11206
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
LTR 11221..11401
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 11448..11573
/label=HIV-1 Psi
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 12066..12299
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 12484..12528
/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 12677..12718
/note="Protein Tat from Human immunodeficiency virus type 1
group M subtype B (isolate WMJ22). Accession#: P12509"
misc_feature 12826..12943
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
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