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pSIN-PAmCherry-KFERQ-NE vector (V017137)

Price Information

Cat No. Plasmid Name Availability Add to cart
V017137 pSIN-PAmCherry-KFERQ-NE 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:
pSIN-PAmCherry-KFERQ-NE
Antibiotic Resistance:
Ampicillin
Length:
8368 bp
Type:
Constitutive expression
Replication origin:
ori
Host:
Mammalian cells, Lentivirus
Selection Marker:
Puro
Promoter:
EF-1α
Growth Strain(s):
Stbl3
Growth Temperature:
37℃

pSIN-PAmCherry-KFERQ-NE vector Map

Copy Sequence View Map
pSIN-PAmCherry-KFERQ-NE8368 bp400800120016002000240028003200360040004400480052005600600064006800720076008000EF-1-alpha promoterEF-1-alpha promoterPAmCherry1IRES2PuroR3' LTR (Delta-U3)bGH poly(A) signalAmpR promoterAmpRori3' LTRHIV-1 PsiRREgp41 peptideProtein TatcPPT/CTS

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.

pSIN-PAmCherry-KFERQ-NE vector Sequence

LOCUS       V017137                 8368 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  Exported.
ACCESSION   V017137
VERSION     V017137
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 8368)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..8368
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..1179
                     /label="EF-1-alpha promoter"
                     /note="strong constitutive promoter for human elongation
                     factor EF-1-alpha"
     CDS             1193..1900
                     /label="PAmCherry1"
                     /note="photoactivatable monomeric derivative of DsRed
                     fluorescent protein (Subach et al., 2009)"
     misc_feature    2083..2661
                     /label="IRES2"
                     /note="internal ribosome entry site (IRES) of the
                     encephalomyocarditis virus (EMCV)"
     CDS             2681..3277
                     /label="PuroR"
                     /note="puromycin N-acetyltransferase"
     LTR             3483..3716
                     /label="3' LTR (Delta-U3)"
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    3873..4097
                     /label="bGH poly(A) signal"
                     /note="bovine growth hormone polyadenylation signal"
     promoter        4193..4297
                     /label="AmpR promoter"
     CDS             4298..5155
                     /label="AmpR"
                     /note="beta-lactamase"
     rep_origin      5329..5917
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     LTR             6076..6709
                     /label="3' LTR"
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    6756..6881
                     /label="HIV-1 Psi"
                     /note="packaging signal of human immunodeficiency virus
                     type 1"
     misc_feature    7374..7607
                     /label="RRE"
                     /note="The Rev response element (RRE) of HIV-1 allows for
                     Rev-dependent mRNA export from the nucleus to the
                     cytoplasm."
     CDS             7792..7836
                     /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             7985..8026
                     /note="Protein Tat from Human immunodeficiency virus type 1
                     group M subtype B (isolate WMJ22). Accession#: P12509"
                     /label="Protein Tat"
     misc_feature    8123..8240
                     /label="cPPT/CTS"
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
     promoter        8368
                     /label="EF-1-alpha promoter"
                     /note="strong constitutive promoter for human elongation
                     factor EF-1-alpha"