Price Information
| Cat No. | Plasmid Name | Availability | Buy one, get one free! (?) |
|---|---|---|---|
| V000058 | pfNL43-dE-EGFP | In stock, instant shipping |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
pfNL43-dE-EGFP can be employed to construct HIV pseudoviruses, enabling visualisation of the infection process through EGFP labelling. This facilitates the detection of compounds' anti-HIV activity and assists in evaluating therapeutic efficacy.
- Vector Name:
- pfNL43-dE-EGFP
- Antibiotic Resistance:
- Ampicillin
- Length:
- 12872 bp
- Type:
- Mammalian Expression, Lentiviral, Retroviral
- Replication origin:
- ori
- Copy Number:
- High Copy
- Growth Strain(s):
- Stbl3
pfNL43-dE-EGFP 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
- Ratanabunyong S, Seetaha S, Hannongbua S, Yanaka S, Yagi-Utsumi M, Kato K, Paemanee A, Choowongkomon K. Biophysical Characterization of Novel DNA Aptamers against K103N/Y181C Double Mutant HIV-1 Reverse Transcriptase. Molecules. 2022 Jan 3;27(1):285. doi: 10.3390/molecules27010285. PMID: 35011517; PMCID: PMC8746315.
pfNL43-dE-EGFP vector Sequence
LOCUS Exported 12872 bp DNA circular SYN 03-NOV-2025
DEFINITION Exported.
ACCESSION V000058
VERSION .
KEYWORDS pfNL43-dE-EGFP
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 12872)
TITLE NLdDIS
JOURNAL Unpublished
REFERENCE 2 (bases 1 to 12872)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 12872)
TITLE Direct Submission
REFERENCE 4 (bases 1 to 12872)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName:
"Unpublished"
COMMENT SGRef: number: 2; type: "Journal Article"
COMMENT SGRef: number: 3; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..12872
/mol_type="other DNA"
/organism="synthetic DNA construct"
LTR 69..326
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 373..498
/label=HIV-1 Psi
/note="packaging signal of human immunodeficiency virus
type 1"
CDS 482..1984
/codon_start=1
/gene="gag"
/product="gag protein from human immunodeficiency virus 1"
/label=HIV-1 gag
/translation="MGARASVLSGGELDKWEKIRLRPGGKKQYKLKHIVWASRELERFA
VNPGLLETSEGCRQILGQLQPSLQTGSEELRSLYNTIAVLYCVHQRIDVKDTKEALDKI
EEEQNKSKKKAQQAAADTGNNSQVSQNYPIVQNLQGQMVHQAISPRTLNAWVKVVEEKA
FSPEVIPMFSALSEGATPQDLNTMLNTVGGHQAAMQMLKETINEEAAEWDRLHPVHAGP
IAPGQMREPRGSDIAGTTSTLQEQIGWMTHNPPIPVGEIYKRWIILGLNKIVRMYSPTS
ILDIRQGPKEPFRDYVDRFYKTLRAEQASQEVKNWMTETLLVQNANPDCKTILKALGPG
ATLEEMMTACQGVGGPGHKARVLAEAMSQVTNPATIMIQKGNFRNQRKTVKCFNCGKEG
HIAKNCRAPRKKGCWKCGKEGHQMKDCTERQANFLGKIWPSHKGRPGNFLQSRPEPTAP
PEESFRFGEETTTPSQKQEPIDKELYPLASLRSLFGSDPSSQ"
CDS 1777..4788
/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
SCDKCQLKGEAMHGQVDCSPGIWQLDCTHLEGKVILVAVHVASGYIEAEVIPAETGQET
AYFLLKLAGRWPVKTVHTDNGSNFTSTTVKAACWWAGIKQEFGIPYNPQSQGVIESMNK
ELKKIIGQVRDQAEHLKTAVQMAVFIHNFKRKGGIGGYSAGERIVDIIATDIQTKELQK
QITKIQNFRVYYRDSRDPVWKGPAKLLWKGEGAVVIQDNSDIKVVPRRKAKIIRDYGKQ
MAGDDCVASRQDED"
misc_feature 4473..4590
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
CDS 4733..5308
/gene="vif"
/label=Virion infectivity factor
/note="Virion infectivity factor from Human
immunodeficiency virus type 1 group M subtype B (isolate
NY5). Accession#: P12504"
CDS 5522..5695
/label=Protein Tat
/note="Protein Tat from Human immunodeficiency virus type 1
group M subtype B (isolate BH5). Accession#: P04612"
CDS 5753..5995
/gene="vpu"
/label=Protein Vpu
/note="Protein Vpu from Human immunodeficiency virus type 1
group M subtype B (isolate BH10). Accession#: P69699"
regulatory 6045..6054
/label=Kozak sequence
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
/regulatory_class="other"
CDS 6051..6767
/label=EGFP
/note="enhanced GFP"
misc_feature 7289..7522
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 7707..7751
/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 8045..8071
/label=NES
/note="nuclear export signal from the HIV Rev protein
(Fischer et al., 1995)"
CDS 8317..8343
/gene="nef"
/label=Protein Nef
/note="Protein Nef from Human immunodeficiency virus type 1
group M subtype D (isolate Z84). Accession#: P12481"
LTR 8606..9239
/label=3' LTR
/note="3' long terminal repeat (LTR) from HIV-1"
primer_bind complement(10235..10252)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(10406..10994)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(11165..12025)
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTKPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
promoter complement(12026..12130)
/label=AmpR promoter
primer_bind 12198..12216
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
primer_bind complement(12254..12276)
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind 12376..12395
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"