LOCUS       sequence-464796-        5431 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express Acceptor Peptide (AP) tagged Ubiquitin lacking the two 
            terminal glycines.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5431)
  AUTHORS   Mukhopadhyay U, Levantovsky S, Carusone TM, Gharbi S, Stein F, 
            Behrends C, Bhogaraju S
  TITLE     A ubiquitin-specific, proximity-based labeling approach for the 
            identification of ubiquitin ligase substrates.
  JOURNAL   Sci Adv. 2024 Aug 9;10(32):eadp3000. doi: 10.1126/sciadv.adp3000. 
            Epub 2024 Aug 9.
  PUBMED    39121224
REFERENCE   2  (bases 1 to 5431)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Sci Adv."; 
            date: "2024-08-9"; volume: "10(32):eadp3000. doi"; pages: " 
            10.1126/sciadv.adp3000. Epub 2024 Aug 9"
COMMENT     Sequence Label: pcDNA5-AP-Ub-dGG
FEATURES             Location/Qualifiers
     source          1..5431
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    500..621
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(537..556)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     591..610
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(670..686)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(670..686)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(683..705)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    694..710
                     /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(join(718..724,725..742,743..748))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    763..784
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(901..918)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1072..1660)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1152..1171)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1831..2622,2623..2691))
                     /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     2454..2473
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2692..2796)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2871..2890)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        3062..3441
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3442..3645
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3596..3616
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    3647..3665
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3668..3686
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     3696..3720
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             3811..3855
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     primer_bind     complement(4210..4227)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4216..4440
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4573..4592)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4724..4771
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             join(4779..5431,1..370)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             join(4779..5431,1..370)
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
ORIGIN
        1 ggcgatgttc ggggattccc aatacgaggt cgccaacatc ttcttctgga ggccgtggtt
       61 ggcttgtatg gagcagcaga cgcgctactt cgagcggagg catccggagc ttgcaggatc
      121 gccgcggctc cgggcgtata tgctccgcat tggtcttgac caactctatc agagcttggt
      181 tgacggcaat ttcgatgatg cagcttgggc gcagggtcga tgcgacgcaa tcgtccgatc
      241 cggagccggg actgtcgggc gtacacaaat cgcccgcaga agcgcggccg tctggaccga
      301 tggctgtgta gaagtactcg ccgatagtgg aaaccgacgc cccagcactc gtccgagggc
      361 aaaggaatag cacgtactac gagatttcga ttccaccgcc gccttctatg aaaggttggg
      421 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
      481 ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa
      541 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
      601 caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc
      661 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
      721 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
      781 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
      841 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
      901 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
      961 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     1021 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     1081 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     1141 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     1201 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     1261 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     1321 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     1381 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     1441 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     1501 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     1561 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     1621 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     1681 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     1741 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     1801 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     1861 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     1921 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     1981 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     2041 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     2101 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     2161 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     2221 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     2281 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     2341 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     2401 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     2461 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     2521 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     2581 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     2641 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     2701 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     2761 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     2821 tgacgtcgac ggatcgggag atctcccgat cccctatggt gcactctcag tacaatctgc
     2881 tctgatgccg catagttaag ccagtatctg ctccctgctt gtgtgttgga ggtcgctgag
     2941 tagtgcgcga gcaaaattta agctacaaca aggcaaggct tgaccgacaa ttgcatgaag
     3001 aatctgctta gggttaggcg ttttgcgctg cttcgcgatg tacgggccag atatacgcgt
     3061 tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt agttcatagc
     3121 ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc
     3181 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg
     3241 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat
     3301 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc
     3361 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta
     3421 ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag
     3481 cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt
     3541 tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa
     3601 atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctctccc tatcagtgat
     3661 agagatctcc ctatcagtga tagagatcgt cgacgagctc gtttagtgaa ccgtcagatc
     3721 gcctggagac gccatccacg ctgttttgac ctccatagaa gacaccggga ccgatccagc
     3781 ctccggactc tagcgtttaa acttaagatg ggcctgaacg acatcttcga ggcccagaag
     3841 atcgagtggc acgagggctc cggctccacc tccggctccg gcatgcagat cttcgtgaag
     3901 actctgactg gtaagaccat caccctcgag gttgagccca gtgacaccat tgagaatgtc
     3961 aaggcaaaga tccaagataa ggaaggcatc cctcctgacc agcagaggct gatctttgct
     4021 ggaaaacagc tggaagatgg gcgcaccctg tctgactaca acatccagaa agagtccacc
     4081 ctgcacctgg tactccgtct cagatagggg taataaggat ccactagtcc agtgtggtgg
     4141 aattctgcag atatccagca cagtggcggc cgctcgagtc tagagggccc gtttaaaccc
     4201 gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg
     4261 tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa
     4321 ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca
     4381 gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg
     4441 cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg
     4501 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
     4561 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     4621 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
     4681 tcgaccccaa aaaacttgat tagggtgatg gttcacgtac ctagaagttc ctattccgaa
     4741 gttcctattc tctagaaagt ataggaactt ccttggccaa aaagcctgaa ctcaccgcga
     4801 cgtctgtcga gaagtttctg atcgaaaagt tcgacagcgt ctccgacctg atgcagctct
     4861 cggagggcga agaatctcgt gctttcagct tcgatgtagg agggcgtgga tatgtcctgc
     4921 gggtaaatag ctgcgccgat ggtttctaca aagatcgtta tgtttatcgg cactttgcat
     4981 cggccgcgct cccgattccg gaagtgcttg acattgggga attcagcgag agcctgacct
     5041 attgcatctc ccgccgtgca cagggtgtca cgttgcaaga cctgcctgaa accgaactgc
     5101 ccgctgttct gcagccggtc gcggaggcca tggatgcgat cgctgcggcc gatcttagcc
     5161 agacgagcgg gttcggccca ttcggaccgc aaggaatcgg tcaatacact acatggcgtg
     5221 atttcatatg cgcgattgct gatccccatg tgtatcactg gcaaactgtg atggacgaca
     5281 ccgtcagtgc gtccgtcgcg caggctctcg atgagctgat gctttgggcc gaggactgcc
     5341 ccgaagtccg gcacctcgtg cacgcggatt tcggctccaa caatgtcctg acggacaatg
     5401 gccgcataac agcggtcatt gactggagcg a
//
