Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V001086 | psPAX2-D64V | 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
psPAX2-D64V is derived from psPAX2 plasmid. It is a packaging plasmid for making Integrase deficient lentiviral vectors since has a point mutation in the integrase gene.
- Vector Name:
- psPAX2-D64V
- Antibiotic Resistance:
- Ampicillin
- Length:
- 10644 bp
- Type:
- integrase deficient lentiviral packaging vector
- Replication origin:
- ori
- Growth Strain(s):
- stbl3
- Growth Temperature:
- 37℃
psPAX2-D64V 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
- Certo MT, Ryu BY, Annis JE, Garibov M, Jarjour J, Rawlings DJ, Scharenberg AM. Tracking genome engineering outcome at individual DNA breakpoints. Nat Methods. 2011 Jul 10;8(8):671-6. doi: 10.1038/nmeth.1648.
psPAX2-D64V vector Sequence
LOCUS Exported 10644 bp ds-DNA circular SYN 13-MAY-2021
DEFINITION psPAX2-D64V is derived from psPAX2 plasmid. It is a packaging
plasmid for making Integrase deficient lentiviral vectors since has
a point mutation in the integrase gene..
ACCESSION .
VERSION .
KEYWORDS psPAX2-D64V
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 10644)
AUTHORS Certo MT, Ryu BY, Annis JE, Garibov M, Jarjour J, Rawlings DJ,
Scharenberg AM
TITLE Tracking genome engineering outcome at individual DNA breakpoints.
JOURNAL Nat Methods. 2011 Jul 10;8(8):671-6. doi: 10.1038/nmeth.1648.
PUBMED 21743461
REFERENCE 2 (bases 1 to 10644)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; doi: "10.1038/nmeth";
journalName: "Nat Methods"; date: "2011-07-10- 10"; volume: "8";
issue: "8"; pages: "671-6"
FEATURES Location/Qualifiers
source 1..10644
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 1..196
/label=SV40 promoter
/note="SV40 early promoter"
rep_origin 47..182
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 109..128
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
polyA_signal 202..336
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(239..258)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 293..312
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(404..421)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(575..1163)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(655..674)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(1334..2194)
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
primer_bind 1957..1976
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(2195..2299)
/gene="bla"
/label=AmpR promoter
enhancer 2330..2709
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
intron 2922..3938
/label=chimeric intron
/note="chimera between introns from chicken beta-actin and
rabbit beta-globin"
primer_bind 3862..3884
/label=pCAGGS-5
/note="Chimeric intron in CAG promoter, forward primer"
primer_bind 3946..3965
/label=pCAG-F
/note="Rabbit beta-globin intron, for pCAG plasmids,
forward primer"
CDS 4044..5546
/codon_start=1
/gene="gag"
/product="gag protein from human immunodeficiency virus 1"
/label=HIV-1 gag
/translation="MGARASVLSGGELDRWEKIRLRPGGKKKYKLKHIVWASRELERFA
VNPGLLETSEGCRQILGQLQPSLQTGSEELRSLYNTVATLYCVHQRIEIKDTKEALDKI
EEEQNKSKKKAQQAAADTGHSNQVSQNYPIVQNIQGQMVHQAISPRTLNAWVKVVEEKA
FSPEVIPMFSALSEGATPQDLNTMLNTVGGHQAAMQMLKETINEEAAEWDRVHPVHAGP
IAPGQMREPRGSDIAGTTSTLQEQIGWMTHNPPIPVGEIYKRWIILGLNKIVRMYSPTS
ILDIRQGPKEPFRDYVDRFYKTLRAEQASQEVKNWMTETLLVQNANPDCKTILKALGPG
ATLEEMMTACQGVGGPGHKARVLAEAMSQVTNPATIMIQKGNFRNQRKTVKCFNCGKEG
HIAKNCRAPRKKGCWKCGKEGHQMKDCTERQANFLGKIWPSHKGRPGNFLQSRPEPTAP
PEESFRFGEETTTPSQKQEPIDKELYPLASLRSLFGSDPSSQ"
CDS 5339..8350
/codon_start=1
/gene="pol"
/product="pol protein from human immunodeficiency virus 1"
/label=HIV-1 pol
/translation="FFREDLAFPQGKAREFSSEQTRANSPTRRELQVWGRDNNSLSEAG
ADRQGTVSFSFPQITLWQRPLVTIKIGGQLKEALLDTGADDTVLEEMNLPGRWKPKMIG
GIGGFIKVRQYDQILIEICGHKAIGTVLVGPTPVNIIGRNLLTQIGCTLNFPISPIETV
PVKLKPGMDGPKVKQWPLTEEKIKALVEICTEMEKEGKISKIGPENPYNTPVFAIKKKD
STKWRKLVDFRELNKRTQDFWEVQLGIPHPAGLKQKKSVTVLDVGDAYFSVPLDKDFRK
YTAFTIPSINNETPGIRYQYNVLPQGWKGSPAIFQCSMTKILEPFRKQNPDIVIYQYMD
DLYVGSDLEIGQHRTKIEELRQHLLRWGFTTPDKKHQKEPPFLWMGYELHPDKWTVQPI
VLPEKDSWTVNDIQKLVGKLNWASQIYAGIKVRQLCKLLRGTKALTEVVPLTEEAELEL
AENREILKEPVHGVYYDPSKDLIAEIQKQGQGQWTYQIYQEPFKNLKTGKYARMKGAHT
NDVKQLTEAVQKIATESIVIWGKTPKFKLPIQKETWEAWWTEYWQATWIPEWEFVNTPP
LVKLWYQLEKEPIIGAETFYVDGAANRETKLGKAGYVTDRGRQKVVPLTDTTNQKTELQ
AIHLALQDSGLEVNIVTDSQYALGIIQAQPDKSESELVSQIIEQLIKKEKVYLAWVPAH
KGIGGNEQVDKLVSAGIRKVLFLDGIDKAQEEHEKYHSNWRAMASDFNLPPVVAKEIVA
SCDKCQLKGEAMHGQVDCSPGIWQLVCTHLEGKVILVAVHVASGYIEAEVIPAETGQET
AYFLLKLAGRWPVKTVHTDNGSNFTSTTVKAACWWAGIKQEFGIPYNPQSQGVIESMNK
ELKKIIGQVRDQAEHLKTAVQMAVFIHNFKRKGGIGGYSAGERIVDIIATDIQTKELQK
QITKIQNFRVYYRDSRDPVWKGPAKLLWKGEGAVVIQDNSDIKVVPRRKAKIIRDYGKQ
MAGDDCVASRQDED"
misc_feature 8035..8152
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
misc_feature 8924..9157
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 9342..9386
/codon_start=1
/product="antigenic peptide corresponding to amino acids
655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik
et al., 2013)"
/label=gp41 peptide
/note="recognized by the 2H10 single-chain llama nanobody"
/translation="KNEQELLELDKWASL"
CDS 9680..9706
/codon_start=1
/product="nuclear export signal from the HIV Rev protein
(Fischer et al., 1995)"
/label=NES
/translation="LPPLERLTL"
primer_bind complement(9991..10010)
/label=Bglob-pA-R
/note="Rabbit beta-globin polyA region, reverse primer"
polyA_signal 10056..10111
/label=beta-globin poly(A) signal
/note="rabbit beta-globin polyadenylation signal (Gil and
Proudfoot, 1987)"
primer_bind complement(10110..10129)
/label=rbglobpA-R
/note="Rabbit beta-globin polyA, reverse primer. Also
called rb-glob-pA-term-R"
primer_bind complement(10472..10488)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(10472..10488)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(10485..10507)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 10496..10512
/label=lac operator
/bound_moiety="lac repressor encoded by lacI"
/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(10520..10550)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 10565..10586
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."