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pMDLg/pRRE vector (V012789)

Price Information

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V012789 pMDLg/pRRE In stock, instant shipping

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Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.

Basic Vector Information

3rd generation lentiviral packaging plasmid; Contains Gag and Pol; also requires pRSV-Rev and envelope expressing plasmid

Vector Name:
pMDLg/pRRE
Antibiotic Resistance:
Ampicillin
Length:
8890 bp
Type:
Viral Expression & Packaging Vectors
Replication origin:
ori
Source/Author:
Dr. Didier Trono's lab
Copy Number:
High copy number
Promoter:
CMV+intron
Growth Strain(s):
Stable
Growth Temperature:
37℃
Expression Method:
Constiutive, Stable

pMDLg/pRRE vector Map

Copy Sequence View Map
pMDLg/pRRE8890 bp40080012001600200024002800320036004000440048005200560060006400680072007600800084008800CMV enhancerCMV promoterbeta-globin intronHIV-1 polRREbeta-globin poly(A)oriAmpRAmpR promoter

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

  • Chen T, Huang D, Chen G, Yang T, Yi J, Tian M. Efficient and sustained IGF-1 expression in the adipose tissue-derived stem cells mediated via a lentiviral vector. J Mol Histol. 2015 Feb;46(1):1-11. doi: 10.1007/s10735-014-9599-7. Epub 2014 Dec 3. PMID: 25467975.

  • Li M, Husic N, Lin Y, Snider BJ. Production of lentiviral vectors for transducing cells from the central nervous system. J Vis Exp. 2012 May 24;(63):e4031. doi: 10.3791/4031. PMID: 22664962; PMCID: PMC3466950.

pMDLg/pRRE vector Sequence

LOCUS       Exported                8890 bp DNA     circular SYN 29-AUG-2024
DEFINITION  synthetic circular DNA.
ACCESSION   .
VERSION     .
KEYWORDS    pMDLg-pRRE
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8890)
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 8890)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 8890)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..8890
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        161..540
                     /note="CMV enhancer"
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        541..744
                     /note="CMV promoter"
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     intron          878..1353
                     /label=beta-globin intron
                     /note="internally truncated intron from human beta-globin"
     CDS             1437..2939
                     /codon_start=1
                     /label=HIV-1 gag
                     /note="HIV-1 gag"
                     /translation="MGARASVLSGGELDRWEKIRLRPGGKKKYKLKHIVWASRELERFA
                     VNPGLLETSEGCRQILGQLQPSLQTGSEELRSLYNTVATLYCVHQRIEIKDTKEALDKI
                     EEEQNKSKKKAQQAAADTGHSNQVSQNYPIVQNIQGQMVHQAISPRTLNAWVKVVEEKA
                     FSPEVIPMFSALSEGATPQDLNTMLNTVGGHQAAMQMLKETINEEAAEWDRVHPVHAGP
                     IAPGQMREPRGSDIAGTTSTLQEQIGWMTHNPPIPVGEIYKRWIILGLNKIVRMYSPTS
                     ILDIRQGPKEPFRDYVDRFYKTLRAEQASQEVKNWMTETLLVQNANPDCKTILKALGPG
                     ATLEEMMTACQGVGGPGHKARVLAEAMSQVTNPATIMIQKGNFRNQRKTVKCFNCGKEG
                     HIAKNCRAPRKKGCWKCGKEGHQMKDCTERQANFLGKIWPSHKGRPGNFLQSRPEPTAP
                     PEESFRFGEETTTPSQKQEPIDKELYPLASLRSLFGSDPSSQ"
     CDS             2732..5740
                     /codon_start=1
                     /label=HIV-1 pol
                     /note="pol protein from human immunodeficiency virus 1"
                     /translation="FFREDLAFPQGKAREFSSEQTRANSPTRRELQVWGRDNNSLSEAG
                     ADRQGTVSFSFPQITLWQRPLVTIKIGGQLKEALLDTGADDTVLEEMNLPGRWKPKMIG
                     GIGGFIKVRQYDQILIEICGHKAIGTVLVGPTPVNIIGRNLLTQIGCTLNFPISPIETV
                     PVKLKPGMDGPKVKQWPLTEEKIKALVEICTEMEKEGKISKIGPENPYNTPVFAIKKKD
                     STKWRKLVDFRELNKRTQDFWEVQLGIPHPAGLKQKKSVTVLDVGDAYFSVPLDKDFRK
                     YTAFTIPSINNETPGIRYQYNVLPQGWKGSPAIFQCSMTKILEPFRKQNPDIVIYQYMD
                     DLYVGSDLEIGQHRTKIEELRQHLLRWGFTTPDKKHQKEPPFLWMGYELHPDKWTVQPI
                     VLPEKDSWTVNDIQKLVGKLNWASQIYAGIKVRQLCKLLRGTKALTEVVPLTEEAELEL
                     AENREILKEPVHGVYYDPSKDLIAEIQKQGQGQWTYQIYQEPFKNLKTGKYARMKGAHT
                     NDVKQLTEAVQKIATESIVIWGKTPKFKLPIQKETWEAWWTEYWQATWIPEWEFVNTPP
                     LVKLWYQLEKEPIIGAETFYVDGAANRETKLGKAGYVTDRGRQKVVPLTDTTNQKTELQ
                     AIHLALQDSGLEVNIVTDSQYALGIIQAQPDKSESELVSQIIEQLIKKEKVYLAWVPAH
                     KGIGGNEQVDKLVSAGIRKVLFLDGIDKAQEEHEKYHSNWRAMASDFNLPPVVAKEIVA
                     SCDKCQLKGEAMHGQVDCSPGIWQLDCTHLEGKVILVAVHVASGYIEAEVIPAETGQET
                     AYFLLKLAGRWPVKTVHTDNGSNFTSTTVKAACWWAGIKQEFGIPYNPQSQGVIESMNK
                     ELKKIIGQVRDQAEHLKTAVQMAVFIHNFKRKGGIGGYSAGERIVDIIATDIQTKELQK
                     QITKIQNFRVYYRDSRDPVWKGPAKLLWKGEGAVVIQDNSDIKVVPRRKAKIIRDYGKQ
                     MAGDDCVASRQDED"
     misc_feature    5768..6001
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     polyA_signal    6232..6626
                     /label=beta-globin poly(A)
                     /note="human beta-globin polyadenylation signal"
     rep_origin      complement(7136..7724)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(7898..8755)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(8756..8860)
                     /label=AmpR promoter