Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V004398 | pHAGE_puro | 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:
- pHAGE_puro
- Antibiotic Resistance:
- Ampicillin
- Length:
- 7585 bp
- Type:
- Lentiviral
- Replication origin:
- ori
- Selection Marker:
- Puromycin
- Copy Number:
- High Copy
- Promoter:
- CMV
- Cloning Method:
- Gibson Cloning
- 5' Primer:
- CMV_F
pHAGE_puro 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.
pHAGE_puro vector Sequence
LOCUS V004398 7585 bp DNA circular SYN 20-DEC-2021
DEFINITION Exported.
ACCESSION V004398
VERSION V004398
KEYWORDS pHAGE_puro
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
.
REFERENCE 1 (bases 1 to 7585)
AUTHORS Lu B, Klingbeil O, Tarumoto Y, Somerville TDD, Huang YH, Wei Y, Wai
DC, Low JKK, Milazzo JP, Wu XS, Cao Z, Yan X, Demerdash OE, Huang G,
Mackay JP, Kinney JB, Shi J, Vakoc CR
TITLE A Transcription Factor Addiction in Leukemia Imposed by the MLL
Promoter Sequence.
JOURNAL Cancer Cell. 2018 Dec 10;34(6):970-981.e8. doi:
10.1016/j.ccell.2018.10.015. Epub 2018 Nov 29.
PUBMED 30503706
REFERENCE 2 (bases 1 to 7585)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 7585)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; doi:
"10.1016/j.ccell.2018.10.015"; journalName: "Cancer Cell"; date:
"2018-12-10- 10"; volume: "34"; issue: "6"; pages: "970-981.e8"
SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..7585
/mol_type="other DNA"
/organism="synthetic DNA construct"
primer_bind 52..71
/label="mPGK-F"
/note="Mouse PGK promoter, forward primer"
CDS 140..736
/label="PuroR"
/note="puromycin N-acetyltransferase"
misc_feature 753..1341
/label="WPRE"
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
LTR 1416..1649
/label="3' LTR (Delta-U3)"
/note="self-inactivating 3' long terminal repeat (LTR) from
HIV-1"
primer_bind complement(1768..1786)
/label="pBRforEco"
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 1854..1958
/label="AmpR promoter"
CDS 1959..2816
/label="AmpR"
/note="beta-lactamase"
rep_origin 2990..3578
/label="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 3732..3749
/label="L4440"
/note="L4440 vector, forward primer"
primer_bind 3841..3860
/label="SV40pro-F"
/note="SV40 promoter/origin, forward primer"
protein_bind 3989..4010
/label="CAP binding site"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 4025..4055
/label="lac promoter"
/note="promoter for the E. coli lac operon"
protein_bind 4063..4079
/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 4087..4103
/label="M13 rev"
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 4153..4172
/label="EBV-rev"
/note="SV40 polyA terminator, reverse primer"
LTR 4364..4997
/label="3' LTR"
/note="3' long terminal repeat (LTR) from HIV-1"
misc_feature 5044..5169
/label="HIV-1 Psi"
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 5666..5899
/label="RRE"
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 6084..6128
/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 6277..6318
/note="Protein Tat from Human immunodeficiency virus type 1
group M subtype B (isolate WMJ22). Accession#: P12509"
/label="Protein Tat"
misc_feature 6421..6538
/label="cPPT/CTS"
/note="central polypurine tract and central termination
sequence of HIV-1"
enhancer 6590..6893
/label="CMV enhancer"
/note="human cytomegalovirus immediate early enhancer"
promoter 6894..7097
/label="CMV promoter"
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 7094..7118
/label="LNCX"
/note="Human CMV promoter, forward primer"
promoter 7205..7585
/label="PGK promoter"
/note="mouse phosphoglycerate kinase 1 promoter"