Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V016676 | Fuw-dCas9-Dnmt3a | 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:
- Fuw-dCas9-Dnmt3a
- Antibiotic Resistance:
- Ampicillin
- Length:
- 16074 bp
- Type:
- RNA interference
- Replication origin:
- ori
- Host:
- Mammalian cells, Lentivirus
- Promoter:
- UbC
- Growth Strain(s):
- Stbl3
- Growth Temperature:
- 37℃
Fuw-dCas9-Dnmt3a 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.
Fuw-dCas9-Dnmt3a vector Sequence
LOCUS V016676 16074 bp DNA circular SYN 01-JAN-1980
DEFINITION Exported.
ACCESSION V016676
VERSION V016676
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
.
REFERENCE 1 (bases 1 to 16074)
AUTHORS .
TITLE Direct Submission
FEATURES Location/Qualifiers
source 1..16074
/mol_type="other DNA"
/organism="synthetic DNA construct"
promoter 393..603
/label="SV40 promoter"
/note="SV40 early promoter"
promoter 659..706
/label="EM7 promoter"
/note="synthetic bacterial promoter"
CDS 725..1096
/label="BleoR"
/note="antibiotic-binding protein"
polyA_signal 1229..1362
/label="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
primer_bind complement(1399..1415)
/label="M13 rev"
/note="common sequencing primer, one of multiple similar
variants"
protein_bind complement(1423..1439)
/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(1447..1477)
/label="lac promoter"
/note="promoter for the E. coli lac operon"
protein_bind complement(1492..1513)
/label="CAP binding site"
/note="CAP binding activates transcription in the presence
of cAMP."
rep_origin complement(1801..2389)
/direction=LEFT
/label="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(2563..3420)
/label="AmpR"
/note="beta-lactamase"
promoter complement(3421..3525)
/label="AmpR promoter"
enhancer 3791..4170
/label="CMV enhancer"
/note="human cytomegalovirus immediate early enhancer"
promoter 4171..4373
/label="CMV promoter"
/note="human cytomegalovirus (CMV) immediate early
promoter"
LTR 4388..4568
/label="5' LTR (truncated)"
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 4615..4740
/label="HIV-1 Psi"
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 5233..5466
/label="RRE"
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 5651..5695
/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 5844..5885
/note="Protein Tat from Human immunodeficiency virus type 1
group M subtype B (isolate WMJ22). Accession#: P12509"
/label="Protein Tat"
misc_feature 5993..6110
/label="cPPT/CTS"
/note="central polypurine tract and central termination
sequence of HIV-1"
promoter 6175..7386
/label="UbC promoter"
/note="human ubiquitin C promoter"
CDS 7437..11540
/label="dCas9"
/note="catalytically dead mutant of the Cas9 endonuclease
from the Streptococcus pyogenes Type II CRISPR/Cas system"
CDS 11577..11597
/label="SV40 NLS"
/note="nuclear localization signal of SV40 (simian virus
40) large T antigen"
CDS 11601..11621
/label="SV40 NLS"
/note="nuclear localization signal of SV40 (simian virus
40) large T antigen"
CDS 11628..14363
/label="DNMT3A"
/note="DNA methyltransferase 3A"
misc_feature 14391..14979
/label="WPRE"
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
LTR 15504..15684
/label="5' LTR (truncated)"
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
polyA_signal 15716..15940
/label="bGH poly(A) signal"
/note="bovine growth hormone polyadenylation signal"