Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V006815 | pLX304 Luciferase-V5 blast | 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 constitutive expression of Luciferase with C-terminal V5 tag.
- Vector Name:
- pLX304 Luciferase-V5 blast
- Antibiotic Resistance:
- Ampicillin
- Length:
- 9375 bp
- Type:
- Mammalian Expression, Lentiviral
- Replication origin:
- ori
- Selection Marker:
- Blasticidin
- Copy Number:
- Low Copy
- Promoter:
- hPGK
- Cloning Method:
- Gateway Cloning
- 5' Primer:
- CMV-F
- 3' Primer:
- WPRE-R
- Growth Strain(s):
- Stbl3
- Growth Temperature:
- 37℃
pLX304 Luciferase-V5 blast 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
- Bajikar SS, Wang CC, Borten MA, Pereira EJ, Atkins KA, Janes KA. Tumor-Suppressor Inactivation of GDF11 Occurs by Precursor Sequestration in Triple-Negative Breast Cancer. Dev Cell. 2017 Nov 20;43(4):418-435.e13.
pLX304 Luciferase-V5 blast vector Sequence
LOCUS 40924_29486 9375 bp DNA circular SYN 13-MAY-2021
DEFINITION Lentiviral vector for constitutive expression of Luciferase with
C-terminal V5 tag.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 9375)
AUTHORS Bajikar SS, Wang CC, Borten MA, Pereira EJ, Atkins KA, Janes KA
TITLE Tumor-Suppressor Inactivation of GDF11 Occurs by Precursor
Sequestration in Triple-Negative Breast Cancer.
JOURNAL Dev Cell. 2017 Nov 20;43(4):418-435.e13. doi:
10.1016/j.devcel.2017.10.027.
PUBMED 29161592
REFERENCE 2 (bases 1 to 9375)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 9375)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; doi:
"10.1016/j.devcel.2017.10"; journalName: "Dev Cell"; date:
"2017-11-20- 20"; volume: "43"; issue: "4"; pages: "418-435.e13"
COMMENT SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..9375
/mol_type="other DNA"
/organism="synthetic DNA construct"
primer_bind 112..131
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 365..382
/label=L4440
/note="L4440 vector, forward primer"
protein_bind 499..520
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 535..565
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 573..589
/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)."
primer_bind 597..613
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
promoter 634..652
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
promoter 680..906
/label=RSV promoter
/note="Rous sarcoma virus enhancer/promoter"
LTR 907..1087
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 1134..1259
/label=HIV-1 Psi
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 1752..1985
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 2170..2214
/codon_start=1
/label=gp41 peptide
/note="antigenic peptide corresponding to amino acids 655
to 669 of the HIV envelope protein gp41 (Lutje Hulsik et
al., 2013)"
/translation="KNEQELLELDKWASL"
promoter 2389..2889
/label=hPGK promoter
/note="human phosphoglycerate kinase 1 promoter"
CDS 2901..3296
/codon_start=1
/label=BSD
/note="blasticidin S deaminase"
/translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
KAIVKDSDGQPTAVGIRELLPSGYVWEG"
misc_feature 3356..3473
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
enhancer 3590..3893
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 3894..4097
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
protein_bind 4120..4144
/gene="mutant version of attB"
/label=attB1
/bound_moiety="BP Clonase(TM)"
/note="recombination site for the Gateway(R) BP reaction"
CDS 4147..5796
/codon_start=1
/label=luciferase
/note="firefly luciferase"
/translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
protein_bind complement(5801..5821)
/label=attB2
/note="core recombination site for the Gateway(R) BP
reaction"
CDS 5824..5865
/codon_start=1
/label=V5 tag
/note="epitope tag from simian virus 5"
/translation="GKPIPNPLLGLDST"
misc_feature 5907..6495
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
LTR 6567..6800
/label=3' LTR (Delta-U3)
/note="self-inactivating 3' long terminal repeat (LTR) from
HIV-1"
polyA_signal 6878..7012
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
rep_origin 7039..7174
/label=SV40 ori
/note="SV40 origin of replication"
promoter complement(7195..7213)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(7223..7239)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
rep_origin 7381..7836
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 7862..7966
/label=AmpR promoter
CDS 7967..8824
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
rep_origin 8998..9375
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"