LOCUS Exported 8025 bp ds-DNA circular SYN 13-MAY-2021
DEFINITION A fusion protein of the SunTag 24xGCN4 v1 peptide array to a
mitochondrial targeting domain and mCherry.
ACCESSION .
VERSION .
KEYWORDS pcDNA4TO-mito-mCherry-24xGCN4_v1
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 8025)
AUTHORS Tanenbaum ME, Gilbert LA, Qi LS, Weissman JS, Vale RD
TITLE A Protein-Tagging System for Signal Amplification in Gene Expression
and Fluorescence Imaging.
JOURNAL Cell. 2014 Oct 8. pii: S0092-8674(14)01227-6. doi:
10.1016/j.cell.2014.09.039.
PUBMED 25307933
REFERENCE 2 (bases 1 to 8025)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Cell. 2014
Oct 8. pii: S0092-8674(14)01227-6. doi: 10.1016/j.cell.2014.09.039."
FEATURES Location/Qualifiers
source 1..8025
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind complement(44..63)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 235..614
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
primer_bind 720..740
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
promoter 725..769
/label=CMVd3 promoter
/note="truncated human cytomegalovirus (CMV) immediate
early promoter that yields strongly reduced expression
relative to the full-length promoter"
protein_bind 771..789
/gene="tetO"
/label=tet operator
/bound_moiety="tetracycline repressor TetR"
/note="bacterial operator O2 for the tetR and tetA genes"
protein_bind 792..810
/gene="tetO"
/label=tet operator
/bound_moiety="tetracycline repressor TetR"
/note="bacterial operator O2 for the tetR and tetA genes"
primer_bind 820..844
/label=LNCX
/note="Human CMV promoter, forward primer"
CDS 1163..1867
/codon_start=1
/product="monomeric derivative of DsRed fluorescent protein
(Shaner et al., 2004)"
/label=mCherry
/note="mammalian codon-optimized"
/translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
GRHSTGGMDELYK"
primer_bind complement(1305..1323)
/label=mCherry-R
/note="mCherry, reverse primer"
primer_bind 1573..1592
/label=mCherry-F
/note="mCherry, forward primer"
CDS 1955..2020
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 2036..2101
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 2117..2182
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 2198..2263
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 2279..2344
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 2360..2425
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 2441..2506
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 2522..2587
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 2603..2668
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 2684..2749
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 2765..2830
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 2846..2911
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
primer_bind 2927..2949
/label=pGEX 5
/note="pGEX vectors, Glutathione-S-transferase, forward
primer"
primer_bind 2950..2972
/label=pGEX 3
/note="pGEX vectors, reverse primer"
CDS 2975..3040
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 3056..3121
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 3137..3202
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 3218..3283
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 3299..3364
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 3380..3445
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 3461..3526
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 3542..3607
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 3623..3688
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 3704..3769
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 3785..3850
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
CDS 3866..3931
/codon_start=1
/product="GCN4 peptide that allows binding to an scFv-GFP
fusion protein (Tanenbaum et al., 2014)"
/label=GCN4_v1
/translation="LLPKNYHLENEVARLKKLVGER"
primer_bind complement(4036..4053)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 4042..4266
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 4312..4740
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(4399..4418)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 4609..4630
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(4749..4769)
/label=pBABE 3
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 4754..5083
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 4934..5069
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 4996..5015
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
promoter 5131..5178
/label=EM7 promoter
/note="synthetic bacterial promoter "
CDS 5197..5571
/codon_start=1
/gene="Sh ble from Streptoalloteichus hindustanus"
/product="antibiotic-binding protein"
/label=BleoR
/note="confers resistance to bleomycin, phleomycin, and
Zeocin(TM)"
/translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
EFALRDPAGNCVHFVAEEQD"
polyA_signal 5701..5822
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(5738..5757)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 5792..5811
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(5871..5887)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(5871..5887)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(5884..5906)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 5895..5911
/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(5919..5949)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 5964..5985
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(6102..6119)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(6273..6858)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(6353..6372)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(7029..7889)
/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 7652..7671
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(7890..7994)
/gene="bla"
/label=AmpR promoter
