Lentiviral-TOP-dGFP-reporter vector (V012444)

Price Information

Cat No. Plasmid Name Availability Add to cart
V012444 Lentiviral-TOP-dGFP-reporter In stock, 1 week for quality controls

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

Basic Vector Information

Vector Name:
Lentiviral-TOP-dGFP-reporter
Antibiotic Resistance:
Ampicillin
Length:
7151 bp
Type:
Mammalian Expression, Lentiviral
Replication origin:
ori
Copy Number:
High Copy
Promoter:
RSV
Cloning Method:
Restriction Enzyme

Lentiviral-TOP-dGFP-reporter vector Map

Lentiviral-TOP-dGFP-reporter7151 bp30060090012001500180021002400270030003300360039004200450048005100540057006000630066006900oriL4440CAP binding sitelac promoterlac operatorM13 revT3 promoterRSV promoter5' LTR (truncated)HIV-1 PsiRREgp41 peptideProtein TatcPPT/CTSKozak sequenced2EGFPWPRE3' LTR (Delta-U3)SV40 poly(A) signalSV40 oriT7 promoterM13 fwdf1 oriAmpR promoterAmpR

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.

Lentiviral-TOP-dGFP-reporter vector Sequence

Copy Sequence

Download GenBank File(.gb)

LOCUS       V012444                 7151 bp    DNA     circular SYN 17-AUG-2021
DEFINITION  Exported.
ACCESSION   V012444
VERSION     V012444
KEYWORDS    Lentiviral-TOP-dGFP-reporter
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 7151)
  AUTHORS   Reya T, Duncan AW, Ailles L, Domen J, Scherer DC, Willert K, Hintz
            L, Nusse R, Weissman IL
  TITLE     A role for Wnt signalling in self-renewal of haematopoietic stem
            cells.
  JOURNAL   Nature. 2003 May 22. 423(6938):409-14.
   PUBMED   12717450
REFERENCE   2  (bases 1 to 7151)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 7151)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nature.
            2003 May 22. 423(6938):409-14."
            SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..7151
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      15..603
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     primer_bind     757..774
                     /label="L4440"
                     /note="L4440 vector, forward primer"
     protein_bind    891..912
                     /label="CAP binding site"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        927..957
                     /label="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    965..981
                     /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     989..1005
                     /label="M13 rev"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     promoter        1026..1044
                     /label="T3 promoter"
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        1072..1298
                     /label="RSV promoter"
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             1299..1479
                     /label="5' LTR (truncated)"
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1526..1651
                     /label="HIV-1 Psi"
                     /note="packaging signal of human immunodeficiency virus
                     type 1"
     misc_feature    2144..2377
                     /label="RRE"
                     /note="The Rev response element (RRE) of HIV-1 allows for
                     Rev-dependent mRNA export from the nucleus to the
                     cytoplasm."
     CDS             2562..2606
                     /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             2755..2796
                     /note="Protein Tat from Human immunodeficiency virus type 1
                     group M subtype B (isolate WMJ22). Accession#: P12509"
                     /label="Protein Tat"
     misc_feature    2873..2990
                     /label="cPPT/CTS"
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
     regulatory      3169..3178
                     /label="Kozak sequence"
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3200..4042
                     /label="d2EGFP"
                     /note="EGFP destabilized by fusion to residues 422-461 of
                     mouse ornithine decarboxylase, giving an in vivo half-life
                     of ~2 hours"
     misc_feature    4066..4654
                     /label="WPRE"
                     /note="woodchuck hepatitis virus posttranscriptional
                     regulatory element"
     LTR             4741..4974
                     /label="3' LTR (Delta-U3)"
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    5046..5180
                     /label="SV40 poly(A) signal"
                     /note="SV40 polyadenylation signal"
     rep_origin      5207..5342
                     /label="SV40 ori"
                     /note="SV40 origin of replication"
     promoter        complement(5363..5381)
                     /label="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5391..5407)
                     /label="M13 fwd"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     rep_origin      5549..6004
                     /label="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        6030..6134
                     /label="AmpR promoter"
     CDS             6135..6992
                     /label="AmpR"
                     /note="beta-lactamase"