Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V010531 | pInducer10-mir-RUP-PheS | 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:
- pInducer10-mir-RUP-PheS
- Antibiotic Resistance:
- Ampicillin
- Length:
- 14567 bp
- Type:
- Mammalian Expression, Lentiviral
- Replication origin:
- ori
- Selection Marker:
- Puromycin
- Promoter:
- UbC
- Cloning Method:
- Restriction Enzyme
- 5' Primer:
- LNCX_primer
pInducer10-mir-RUP-PheS 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.
pInducer10-mir-RUP-PheS vector Sequence
LOCUS V010531 14567 bp DNA circular SYN 13-MAY-2021
DEFINITION Exported.
ACCESSION V010531
VERSION V010531
KEYWORDS pInducer10-mir-RUP-PheS
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
.
REFERENCE 1 (bases 1 to 14567)
AUTHORS Meerbrey KL, Hu G, Kessler JD, Roarty K, Li MZ, Fang JE,
Herschkowitz JI, Burrows AE, Ciccia A, Sun T, Schmitt EM, Bernardi
RJ, Fu X, Bland CS, Cooper TA, Schiff R, Rosen JM, Westbrook TF,
Elledge SJ
TITLE The pINDUCER lentiviral toolkit for inducible RNA interference in
vitro and in vivo.
JOURNAL Proc Natl Acad Sci U S A. 2011 Mar 1;108(9):3665-70. doi:
10.1073/pnas.1019736108. Epub 2011 Feb 9.
PUBMED 21307310
REFERENCE 2 (bases 1 to 14567)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 14567)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; doi:
"10.1073/pnas.1019736108"; journalName: "Proc Natl Acad Sci U S A";
date: "2011-03-1- 1"; volume: "108"; issue: "9"; pages: "3665-70"
SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..14567
/mol_type="other DNA"
/organism="synthetic DNA construct"
misc_feature 497..730
/label="RRE"
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 915..959
/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 1108..1149
/note="Protein Tat from Human immunodeficiency virus type 1
group M subtype B (isolate WMJ22). Accession#: P12509"
/label="Protein Tat"
misc_feature 1257..1374
/label="cPPT/CTS"
/note="central polypurine tract and central termination
sequence of HIV-1"
CDS complement(1442..1813)
/label="BleoR"
/note="antibiotic-binding protein"
promoter complement(1833..1880)
/label="EM7 promoter"
/note="synthetic bacterial promoter"
protein_bind 1950..2220
/label="tetracycline response element"
/note="contains seven copies of the tetracycline operator
tetO"
promoter 2253..2291
/label="minimal CMV promoter"
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 2288..2312
/label="LNCX"
/note="Human CMV promoter, forward primer"
CDS 2388..3080
/label="TurboRFP"
/note="red fluorescent protein from Entacmaea quadricolor"
ncRNA 3134..3227
/label="5' miR-30a"
/note="sequence upstream of the 71-nt precursor of the
human miR-30a microRNA (Zeng et al., 2002)"
primer_bind 3227..3243
/label="pBluescriptKS"
/note="For pBluescript vector"
primer_bind 3228..3244
/label="KS primer"
/note="common sequencing primer, one of multiple similar
variants"
CDS 3265..3282
/label="thrombin site"
/note="thrombin recognition and cleavage site"
CDS 3283..3306
/label="HRV 3C site"
/note="recognition and cleavage site for human rhinovirus
3C and PreScission proteases"
regulatory 3316..3325
/label="Kozak sequence"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
/regulatory_class="other"
CDS complement(3483..4463)
/gene="pheS"
/label="Phenylalanine--tRNA ligase alpha subunit"
/note="Phenylalanine--tRNA ligase alpha subunit from
Escherichia coli O139:H28 (strain E24377A / ETEC).
Accession#: A7ZMI2"
CDS 4601..4621
/label="TEV site"
/note="tobacco etch virus (TEV) protease recognition and
cleavage site"
ncRNA 4645..4757
/label="3' miR-30a"
/note="sequence downstream of the 71-nt precursor of the
human miR-30a microRNA (Zeng et al., 2002)"
promoter 4808..6019
/label="UbC promoter"
/note="human ubiquitin C promoter"
protein_bind 6026..6059
/label="loxP"
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (ATGTATGC) (Shaw et al., 2021)."
CDS 6066..6809
/label="rtTA-Advanced"
/note="improved tetracycline-controlled transactivator"
protein_bind complement(6813..6846)
/label="loxP"
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (ATGTATGC) (Shaw et al., 2021)."
misc_feature 6862..7444
/label="IRES2"
/note="internal ribosome entry site (IRES) of the
encephalomyocarditis virus (EMCV)"
CDS 7443..8036
/label="PuroR"
/note="puromycin N-acetyltransferase"
misc_feature 8061..8649
/label="WPRE"
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
LTR 8856..9089
/label="3' LTR (Delta-U3)"
/note="self-inactivating 3' long terminal repeat (LTR) from
HIV-1"
polyA_signal 9196..9420
/label="bGH poly(A) signal"
/note="bovine growth hormone polyadenylation signal"
rep_origin 9466..9894
/label="f1 ori"
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(9903..9923)
/label="pBABE 3'"
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 10019..10229
/label="SV40 promoter"
/note="SV40 early promoter"
polyA_signal 11060..11181
/label="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
primer_bind complement(11230..11246)
/label="M13 rev"
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(11230..11246)
/label="M13 Reverse"
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(11243..11265)
/label="M13/pUC Reverse"
/note="In lacZ gene"
protein_bind 11254..11270
/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(11278..11308)
/label="lac promoter"
/note="promoter for the E. coli lac operon"
protein_bind complement(11323..11344)
/label="CAP binding site"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(11461..11478)
/label="L4440"
/note="L4440 vector, forward primer"
rep_origin complement(11632..12220)
/direction=LEFT
/label="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(12394..13251)
/label="AmpR"
/note="beta-lactamase"
promoter complement(13252..13356)
/label="AmpR promoter"
polyA_signal 13404..13538
/label="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
LTR 13706..14339
/label="3' LTR"
/note="3' long terminal repeat (LTR) from HIV-1"