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NGFR vector (V004304)

Price Information

Cat No. Plasmid Name Availability Add to cart
V004304 NGFR 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:
NGFR
Antibiotic Resistance:
Ampicillin
Length:
6522 bp
Type:
Mammalian Expression, Retroviral
Replication origin:
ori
Copy Number:
High Copy
Promoter:
MSCV
Cloning Method:
Restriction Enzyme
5' Primer:
pLXSN 5'

NGFR vector Map

Copy Sequence View Map
NGFR6522 bp300600900120015001800210024002700300033003600390042004500480051005400570060006300oriAmpRAmpR promoter5' LTRMESV Psigag (truncated)IRES23' LTRlac operatorlac promoterCAP binding site

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.

NGFR vector Sequence

LOCUS       40924_2204        6522 bp DNA     circular SYN 08-JUN-2021
DEFINITION  synthetic circular DNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6522)
  AUTHORS   Izon DJ, Punt JA, Xu L, Karnell FG, Allman D, Myung PS, Boerth NJ, 
            Pui JC, Koretzky GA, Pear WS
  TITLE     Notch1 regulates maturation of CD4+ and CD8+ thymocytes by 
            modulating TCR signal strength.
  JOURNAL   Immunity. 2001 Mar . 14(3):253-64.
  PUBMED    11290335
REFERENCE   2  (bases 1 to 6522)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 6522)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Immunity. 
            2001 Mar . 14(3):253-64."
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6522
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(684..1541)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(1542..1646)
                     /label=AmpR promoter
     LTR             2474..2990
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem
                     cell virus"
     misc_feature    3054..3395
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell
                     virus"
     CDS             3462..3878
                     /codon_start=1
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag
                     gene lacking the start codon"
                     /translation="GQTVTTPLSLTLGHWKDVERIAHNQSVDVKKRRWVTFCSAEWPTF
                     NVGWPRDGTFNRDLITQVKIKVFSPGPHGHPDQVPYIVTWEALAFDPPPWVKPFVHPKP
                     PPPLPPSAPSLPLEPPRSTPPRSSLYPALTPSLGA"
     misc_feature    3911..4487
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     LTR             5390..5904
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem
                     cell virus"
     protein_bind    complement(6066..6082)
                     /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)."
     promoter        complement(6090..6120)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(6135..6156)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(join(6444..6522,1..510))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"