Basic Vector Information
- Vector Name:
- pLV3-U6-MCS-sgRNA-EF1a-dCas9-VPR-EGFP
- Antibiotic Resistance:
- Ampicillin
- Length:
- 12451 bp
- Type:
- Genome editing
- Replication origin:
- ori
- Host:
- Mammalian cells, Lentivirus
- Promoter:
- U6
- Growth Temperature:
- 37℃
pLV3-U6-MCS-sgRNA-EF1a-dCas9-VPR-EGFP 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-sgRNA-EF1a-dCas9-VPR-EGFP vector Sequence
LOCUS 62056_15665 12451 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 12451)
AUTHORS .
TITLE Direct Submission
FEATURES Location/Qualifiers
source 1..12451
/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 266..341
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
promoter 403..614
/label=EF-1-alpha core promoter
/note="core promoter for human elongation factor
EF-1-alpha"
CDS 646..666
/label=SV40 NLS
/note="nuclear localization signal of SV40 (simian virus
40) large T antigen"
CDS 676..2391
/label=Cas9(N)
/note="N-terminal portion of Streptococcus pyogenes Cas9
(Zetsche et al., 2015)"
CDS 2965..2985
/label=TEV site
/note="tobacco etch virus (TEV) protease recognition and
cleavage site"
CDS 5134..5154
/label=SV40 NLS
/note="nuclear localization signal of SV40 (simian virus
40) large T antigen"
CDS 6361..7077
/label=EGFP
/note="enhanced GFP"
LTR 7152..7385
/label=3' LTR (Delta-U3)
/note="self-inactivating 3' long terminal repeat (LTR) from
HIV-1"
polyA_signal 7417..7641
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
promoter complement(7738..7768)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind complement(7783..7804)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
rep_origin complement(8092..8680)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(8854..9711)
/label=AmpR
/note="beta-lactamase"
promoter complement(9712..9816)
/label=AmpR promoter
enhancer 10082..10461
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 10462..10664
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
LTR 10679..10859
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 10906..11031
/label=HIV-1 Psi
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 11524..11757
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 11942..11986
/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 12135..12176
/note="Protein Tat from Human immunodeficiency virus type 1
group M subtype B (isolate WMJ22). Accession#: P12509"
misc_feature 12284..12401
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
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