Human ZDHHC6 ORF clone (NM_022494) (Cat. No.: V044508)
Note: It expresses human ZDHHC6, a palmitoyltransferase for protein modification, with a 3xFLAG tag and CopGFP reporter driven by CMV promoter, and PuroR via T2A. It includes WPRE and viral elements (LTRs, HIV-1 Ψ, RRE, cPPT/CTS) for packaging, plus AmpR and ori for bacteria.
- Name:
- pLV3-CMV-ZDHHC6(human)-3xFLAG-CopGFP-Puro
- Accession ID:
- NM_022494.3
- Antibiotic Resistance:
- Ampicillin
- Length:
- 9531 bp
- Type:
- Protein expression
- Replication origin:
- ori
- Host:
- Mammalian cells, Lentivirus
- Selection Marker:
- Puro;CopGFP
- Copy Number:
- High
- Promoter:
- CMV
- 5' Primer:
- CMV-F
- Fusion Tag:
- 3xFLAG
- Growth Strain(s):
- Stbl3
- Growth Temperature:
- 37℃
- Expression Method:
- Constiutive, Stable / Transient
- Gene Synonyms:
- DHHC-6; DHHC6; ZNF376
- Transcript Definition:
- Homo sapiens zinc finger DHHC-type palmitoyltransferase 6 (ZDHHC6), transcript variant 1, mRNA
Resources
Plasmid Protocol
1. Centrifuge at 5,000×g for 5 min.
2. Carefully open the tube and add sterile water to dissolve the DNA: add 20 μl for 5 μg plasmid, and 100 μl for 100 μg plasmid.
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.
Human ZDHHC6 ORF clone (NM_022494) (Cat. No.: V044508) Sequence
LOCUS pLV3-CMV-ZDHHC6( 9531 bp DNA circular SYN 15-OCT-2026
DEFINITION synthetic circular DNA.
ACCESSION V044508
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 9531)
AUTHORS .
TITLE Direct Submission
FEATURES Location/Qualifiers
source 1..9531
/mol_type="other DNA"
/organism="synthetic DNA construct"
LTR 233..413
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 460..585
/label=HIV-1 Psi
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 1078..1311
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 1496..1540
/codon_start=1
/product="antigenic peptide corresponding to amino acids
655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik
et al., 2013)"
/label=gp41 peptide
/note="recognized by the 2H10 single-chain llama nanobody"
/translation="KNEQELLELDKWASL"
misc_feature 1807..1923
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
promoter 2013..2216
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
CDS 2319..3557
/gene="ZDHHC6"
/label=ZDHHC6(NM_022494)
/note="ZDHHC6(NM_022494)"
CDS 3564..3629
/codon_start=1
/product="three tandem FLAG(R) epitope tags, followed by an
enterokinase cleavage site"
/label=3xFLAG
/translation="DYKDHDGDYKDHDIDYKDDDDK"
promoter 3673..3884
/label=EF-1-alpha core promoter
/note="core promoter for human elongation factor
EF-1-alpha"
LTR 3897..4165
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from human
T-cell leukemia virus (HTLV) type 1"
regulatory 4194..4203
/label=Kozak sequence
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
/regulatory_class="other"
CDS 4224..4886
/codon_start=1
/product="green fluorescent protein 2 from Pontellina
plumata, also known as ppluGFP2 (Shagin et al., 2004)"
/label=CopGFP
/translation="PAMEIECRITGTLNGVEFELVGGGEGTPKQGRMTNKMKSTKGALT
FSPYLLSHVMGYGFYHFGTYPSGYENPFLHAINNGGYTNTRIEKYEDGGVLHVSFSYRY
EAGRVIGDFKVVGTGFPEDSVIFTDKIIRSNATVEHLHPMGDNVLVGSFARTFSLRDGG
YYSFVVDSHMHFKSAIHPSILQNGGPMFAFRRVEELHSNTELGIVEYQHAFKTPIAFA"
CDS 4956..5009
/codon_start=1
/product="2A peptide from Thosea asigna virus capsid
protein"
/label=T2A
/note="Eukaryotic ribosomes fail to insert a peptide bond
between the Gly and Pro residues, yielding separate
polypeptides."
/translation="EGRGSLLTCGDVEENPGP"
CDS 5010..5609
/codon_start=1
/gene="pac from Streptomyces alboniger"
/product="puromycin N-acetyltransferase"
/label=PuroR
/note="confers resistance to puromycin"
/translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
misc_feature 5610..6198
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
LTR 6272..6505
/label=3' LTR (Delta-U3)
/note="self-inactivating 3' long terminal repeat (LTR)
from HIV-1"
polyA_signal 6577..6698
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
rep_origin 6738..6873
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind complement(6906..6922)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind 6930..6946
/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(6954..6984)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 6999..7020
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
rep_origin complement(7308..7896)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(8067..8927)
/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"
promoter complement(8928..9032)
/gene="bla"
/label=AmpR promoter
primer_bind 9506..9522
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"