Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V018101 | HA-NEDD8 | In stock, instant shipping |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
The plasmid HA-NEDD8 is designed to express HA-tagged NEDD8, a ubiquitin-like protein critical for post-translational modifications.
- Vector Name:
- HA-NEDD8
- Antibiotic Resistance:
- Ampicillin, 100 μg/mL
- Length:
- 5672 bp
- Type:
- Mammalian Expression Vectors
- Source/Author:
- Edward Yeh
- Selection Marker:
- Neomycin (select with G418)
- Copy Number:
- High copy number
- Promoter:
- CMV
- Fusion Tag:
- HA-tag
- Growth Strain(s):
- Top10
- Growth Temperature:
- 37℃
HA-NEDD8 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.
References
- Kamitani T, Kito K, Nguyen HP, Yeh ET. Characterization of NEDD8, a developmentally down-regulated ubiquitin-like protein. J Biol Chem. 1997;272(45):28557-28562. doi:10.1074/jbc.272.45.28557
HA-NEDD8 vector Sequence
LOCUS Exported 5672 bp DNA circular SYN 10-MAR-2025
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS HA-NEDD8
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5672)
AUTHORS Kamitani T, Kito K, Nguyen HP, Yeh ET
TITLE Characterization of NEDD8, a developmentally down-regulated
ubiquitin-like protein.
JOURNAL J Biol Chem. 1997 Nov 7. 272(45):28557-62.
PUBMED 9353319
REFERENCE 2 (bases 1 to 5672)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "J Biol
Chem. 1997 Nov 7. 272(45):28557-62."
FEATURES Location/Qualifiers
source 1..5672
/mol_type="other DNA"
/organism="synthetic DNA construct"
primer_bind complement(82..102)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 87..416
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 267..402
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 329..348
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
CDS 483..1277
/codon_start=1
/gene="aph(3')-II (or nptII)"
/product="aminoglycoside phosphotransferase from Tn5"
/label=NeoR/KanR
/note="confers resistance to neomycin, kanamycin, and G418
(Geneticin(R))"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
primer_bind complement(537..556)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 1147..1166
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
polyA_signal 1451..1572
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(1488..1507)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 1542..1561
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(1621..1637)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(1621..1637)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(1634..1656)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 1645..1661
/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(1669..1699)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 1714..1735
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(1852..1869)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(2023..2611)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(2103..2122)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(2782..3642)
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
primer_bind 3405..3424
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(3643..3747)
/gene="bla"
/label=AmpR promoter
primer_bind complement(3822..3841)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 4013..4392
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 4393..4596
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 4547..4567
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
primer_bind 4641..4660
/label=T7
/note="T7 promoter, forward primer"
promoter 4641..4659
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
CDS 4676..4702
/codon_start=1
/product="HA (human influenza hemagglutinin) epitope tag"
/label=HA
/translation="YPYDVPDYA"
promoter complement(5003..5021)
/label=SP6 promoter
/note="promoter for bacteriophage SP6 RNA polymerase"
primer_bind complement(5004..5021)
/label=SP6
/note="SP6 promoter, forward primer"
primer_bind complement(5041..5058)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 5047..5271
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin join(5317..5672,1..73)
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(5404..5423)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 5614..5635
/label=F1ori-F
/note="F1 origin, forward primer"