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pRetroX-TRE3G vector (V010376) Gene synthesis in pRetroX-TRE3G backbone

Price Information

Cat No. Plasmid Name Availability Buy one, get one free! (?)
V010376 pRetroX-TRE3G In stock, instant shipping

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

Basic Vector Information

Doxycycline-Inducible Retroviral Vector for Controlled Gene Expression with Tet-On 3G System

Vector Name:
pRetroX-TRE3G
Antibiotic Resistance:
Ampicillin
Length:
6600 bp
Type:
Viral Expression & Packaging Vectors
Replication origin:
ori
Source/Author:
Clontech
Copy Number:
High copy number
Promoter:
mPGK
Growth Strain(s):
Top10
Growth Temperature:
37℃

pRetroX-TRE3G vector Map

Copy Sequence View Map
pRetroX-TRE3G6600 bp300600900120015001800210024002700300033003600390042004500480051005400570060006300CMV enhancerCMV promoter5' LTR (truncated)MMLV Psigag (truncated)TRE3GV promoterMCSPGK promoterPuroR3' LTR (Delta-U3)SV40 promoteroriAmpRAmpR promoterCMV enhancer

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

  • Yang X, Wu R, Shan W, Yu L, Xue B, Shi H. DNA Methylation Biphasically Regulates 3T3-L1 Preadipocyte Differentiation. Mol Endocrinol. 2016 Jun;30(6):677-87. doi: 10.1210/me.2015-1135. Epub 2016 May 4. PMID: 27144289; PMCID: PMC4884341.

pRetroX-TRE3G vector Sequence

LOCUS       pRetroX-TRE3G.        6600 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Retroviral vector for doxycycline-inducible expression of a gene in 
            cells containing the Tet-On(R) 3G transactivator protein.
ACCESSION   .
VERSION     .
KEYWORDS    pRetroX-TRE3G
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6600)
  AUTHORS   Clontech
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 6600)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6600
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        5..309
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        310..513
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     LTR             514..689
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    752..951
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus
                     (MMLV)"
     CDS             1152..1568
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag
                     gene lacking the start codon"
     promoter        1608..1957
                     /label=TRE3GV promoter
                     /note="3rd-generation Tet-responsive promoter that can be 
                     activated by binding of Tet-On(R) 3G, optimized for 
                     retroviral and lentiviral vectors"
     misc_feature    1958..2001
                     /label=MCS
                     /note="MCS"
                     /note="multiple cloning site"
     promoter        2006..2505
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     CDS             2526..3122
                     /label=PuroR
                     /note="puromycin N-acetyltransferase"
     LTR             3306..3731
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     promoter        3997..4326
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      complement(4619..5207)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(5381..6238)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(6239..6343)
                     /label=AmpR promoter
     enhancer        6529..6600
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"