Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V012823 | pHR_Gal4UAS_Pembrolizumab_heavychain_T2A_Pembrolizumab_lightchain_PGK_mCherry | In stock, instant shipping |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
Lentiviral vector for Gal4 inducible leucinezipper Pembrolizumab and constitutive mCherry expression
- Vector Name:
- pHR_Gal4UAS_Pembrolizumab_heavychain_T2A_Pembrolizumab_lightchain_PGK_mCherry
- Antibiotic Resistance:
- Ampicillin
- Length:
- 12269 bp
- Type:
- Viral Expression & Packaging Vectors
- Replication origin:
- ori
- Source/Author:
- Wendell Lim
- Promoter:
- mPGK
- Growth Strain(s):
- stbl3
- Growth Temperature:
- 37℃
pHR_Gal4UAS_Pembrolizumab_heavychain_T2A_Pembrolizumab_lightchain_PGK_mCherry 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.
References
- Roybal KT, Williams JZ, Morsut L, Rupp LJ, Kolinko I, Choe JH, Walker WJ, McNally KA, Lim WA. Engineering T Cells with Customized Therapeutic Response Programs Using Synthetic Notch Receptors. Cell. 2016 Oct 6;167(2):419-432.e16. doi: 10.1016/j.cell.2016.09.011. Epub 2016 Sep 29.
pHR_Gal4UAS_Pembrolizumab_heavychain_T2A_Pembrolizumab_lightchain_PGK_mCherry vector Sequence
LOCUS V012823 12269 bp DNA circular SYN 13-MAY-2021
DEFINITION Exported.
ACCESSION V012823
VERSION V012823
KEYWORDS pHR_Gal4UAS_Pembrolizumab_heavychain_T2A_ Pembrolizumab_lightc
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
.
REFERENCE 1 (bases 1 to 12269)
AUTHORS Roybal KT, Williams JZ, Morsut L, Rupp LJ, Kolinko I, Choe JH,
Walker WJ, McNally KA, Lim WA
TITLE Engineering T Cells with Customized Therapeutic Response Programs
Using Synthetic Notch Receptors.
JOURNAL Cell. 2016 Oct 6;167(2):419-432.e16. doi:
10.1016/j.cell.2016.09.011. Epub 2016 Sep 29.
PUBMED 27693353
REFERENCE 2 (bases 1 to 12269)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; doi:
"10.1016/j.cell.2016.09.011"; journalName: "Cell"; date: "2016-10-6-
6"; volume: "167"; issue: "2"; pages: "419-432.e16"
FEATURES Location/Qualifiers
source 1..12269
/mol_type="other DNA"
/organism="synthetic DNA construct"
CDS 58..87
/label="Myc"
/note="Myc (human c-Myc proto-oncogene) epitope tag"
CDS 1453..1506
/label="T2A"
/note="2A peptide from Thosea asigna virus capsid protein"
CDS 1564..1590
/label="HA"
/note="HA (human influenza hemagglutinin) epitope tag"
CDS 1927..2244
/label="hIg-kappa-CL"
/note="Human immunoglobulin kappa light chain constant
region"
promoter 2370..2869
/label="PGK promoter"
/note="mouse phosphoglycerate kinase 1 promoter"
CDS 2890..3597
/label="mCherry"
/note="monomeric derivative of DsRed fluorescent protein
(Shaner et al., 2004)"
misc_feature 3677..4265
/label="WPRE"
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
LTR 4328..4561
/label="3' LTR (Delta-U3)"
/note="self-inactivating 3' long terminal repeat (LTR) from
HIV-1"
promoter complement(4586..4604)
/label="SP6 promoter"
/note="promoter for bacteriophage SP6 RNA polymerase"
primer_bind complement(4894..4913)
/label="pBRrevBam"
/note="pBR322 vectors, tet region, downstream of BamHI,
reverse primer"
primer_bind complement(5127..5146)
/label="pRS-marker"
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 5246..5268
/label="pGEX 3'"
/note="pGEX vectors, reverse primer"
primer_bind complement(5306..5324)
/label="pBRforEco"
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 5392..5496
/label="AmpR promoter"
CDS 5497..6354
/label="AmpR"
/note="beta-lactamase"
rep_origin 6528..7116
/label="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 7270..7287
/label="L4440"
/note="L4440 vector, forward primer"
promoter 7362..7691
/label="SV40 promoter"
/note="SV40 enhancer and early promoter"
intron 8825..8890
/label="small t intron"
/note="SV40 (simian virus 40) small t antigen intron"
CDS 9020..9040
/label="SV40 NLS"
/note="nuclear localization signal of SV40 (simian virus
40) large T antigen"
polyA_signal 9465..9599
/label="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
LTR 10143..10400
/label="5' LTR (truncated)"
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 10447..10572
/label="HIV-1 Psi"
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 11069..11302
/label="RRE"
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 11487..11531
/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 11680..11721
/note="Protein Tat from Human immunodeficiency virus type 1
group M subtype B (isolate WMJ22). Accession#: P12509"
/label="Protein Tat"
misc_feature 11816..11933
/label="cPPT/CTS"
/note="central polypurine tract and central termination
sequence of HIV-1"
primer_bind 12159..12183
/label="LNCX"
/note="Human CMV promoter, forward primer"