Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V001146 | pSLIK-IDH1-R132H-FLAG | In stock, 1 week for quality controls |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
- Vector Name:
- pSLIK-IDH1-R132H-FLAG
- Antibiotic Resistance:
- Ampicillin
- Length:
- 13969 bp
- Type:
- Mammalian Expression, Lentiviral
- Replication origin:
- ori
- Selection Marker:
- Hygromycin
- Promoter:
- UbC
- Cloning Method:
- Gateway Cloning
- 5' Primer:
- CMV-F
- 3' Primer:
- huB-Cpro-R
pSLIK-IDH1-R132H-FLAG vector Map
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.
pSLIK-IDH1-R132H-FLAG vector Sequence
LOCUS V001146 13969 bp DNA circular SYN 13-MAY-2021
DEFINITION Exported.
ACCESSION V001146
VERSION V001146
KEYWORDS pSLIK-IDH1-R132H-FLAG
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
.
REFERENCE 1 (bases 1 to 13969)
AUTHORS Lewis CA, Parker SJ, Fiske BP, McCloskey D, Gui DY, Green CR, Vokes
NI, Feist AM, Vander Heiden MG, Metallo CM
TITLE Tracing compartmentalized NADPH metabolism in the cytosol and
mitochondria of mammalian cells.
JOURNAL Mol Cell. 2014 Jul 17;55(2):253-63. doi:
10.1016/j.molcel.2014.05.008. Epub 2014 May 29.
PUBMED 24882210
REFERENCE 2 (bases 1 to 13969)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 13969)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; doi:
"10.1016/j.molcel.2014.05.008"; journalName: "Mol Cell"; date:
"2014-07-17- 17"; volume: "55"; issue: "2"; pages: "253-63"
SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..13969
/mol_type="other DNA"
/organism="synthetic DNA construct"
primer_bind complement(11..28)
/label="L4440"
/note="L4440 vector, forward primer"
rep_origin complement(182..770)
/direction=LEFT
/label="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(944..1801)
/label="AmpR"
/note="beta-lactamase"
promoter complement(1802..1906)
/label="AmpR promoter"
primer_bind complement(1981..2000)
/label="pRS-marker"
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 2172..2551
/label="CMV enhancer"
/note="human cytomegalovirus immediate early enhancer"
promoter 2552..2754
/label="CMV promoter"
/note="human cytomegalovirus (CMV) immediate early
promoter"
LTR 2769..2949
/label="5' LTR (truncated)"
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 2996..3121
/label="HIV-1 Psi"
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 3614..3847
/label="RRE"
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 4032..4076
/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 4225..4266
/note="Protein Tat from Human immunodeficiency virus type 1
group M subtype B (isolate WMJ22). Accession#: P12509"
/label="Protein Tat"
misc_feature 4374..4491
/label="cPPT/CTS"
/note="central polypurine tract and central termination
sequence of HIV-1"
protein_bind 4548..4572
/label="attB1"
/note="recombination site for the Gateway(R) BP reaction"
promoter 4604..4918
/label="tight TRE promoter"
/note="Tet-responsive promoter PTight, consisting of seven
tet operator sequences followed by the minimal CMV
promoter"
CDS 4931..6172
/gene="IDH1"
/label="Isocitrate dehydrogenase [NADP] cytoplasmic"
/note="Isocitrate dehydrogenase [NADP] cytoplasmic from
Homo sapiens. Accession#: O75874"
CDS 6194..6259
/label="3xFLAG"
/note="three tandem FLAG(R) epitope tags, followed by an
enterokinase cleavage site"
protein_bind complement(6280..6304)
/label="attB2"
/note="recombination site for the Gateway(R) BP reaction"
promoter 6354..7565
/label="UbC promoter"
/note="human ubiquitin C promoter"
CDS 7690..8433
/label="rtTA-Advanced"
/note="improved tetracycline-controlled transactivator"
misc_feature 8474..9047
/label="IRES"
/note="internal ribosome entry site (IRES) of the
encephalomyocarditis virus (EMCV)"
CDS 9106..10122
/label="HygR"
/note="aminoglycoside phosphotransferase from E. coli"
misc_feature 10591..11179
/label="WPRE"
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
primer_bind complement(11182..11198)
/label="KS primer"
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(11183..11199)
/label="pBluescriptKS"
/note="For pBluescript vector"
LTR 11708..11888
/label="5' LTR (truncated)"
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
polyA_signal 11920..12144
/label="bGH poly(A) signal"
/note="bovine growth hormone polyadenylation signal"
rep_origin 12190..12618
/label="f1 ori"
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 12632..12961
/label="SV40 promoter"
/note="SV40 enhancer and early promoter"
promoter 13009..13056
/label="EM7 promoter"
/note="synthetic bacterial promoter"
CDS 13075..13446
/label="BleoR"
/note="antibiotic-binding protein"
polyA_signal 13579..13712
/label="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
primer_bind complement(13749..13765)
/label="M13 rev"
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(13749..13765)
/label="M13 Reverse"
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(13762..13784)
/label="M13/pUC Reverse"
/note="In lacZ gene"
protein_bind 13773..13789
/label="lac operator"
/bound_moiety="lac repressor encoded by lacI"
/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(13797..13827)
/label="lac promoter"
/note="promoter for the E. coli lac operon"
protein_bind complement(13842..13863)
/label="CAP binding site"
/note="CAP binding activates transcription in the presence
of cAMP."