LOCUS       sequence-465034-        5431 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express Ubiquitin tagged with a modified Acceptor Peptide (AP) .
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-(-2)AP-Ub
FEATURES             Location/Qualifiers
     source          1..5431
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(158..177)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        349..728
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        729..932
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     883..903
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    934..952
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    955..973
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     983..1007
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             1170..1397
                     /codon_start=1
                     /product="human ubiquitin"
                     /label=ubiquitin
                     /translation="MQIFVKTLTGKTITLEVEPSDTIENVKAKIQDKEGIPPDQQRLIF
                     AGKQLEDGRTLSDYNIQKESTLHLVLRLRGG"
     primer_bind     complement(1497..1514)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1503..1727
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(1860..1879)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2011..2058
                     /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             2066..3088
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             2066..3088
                     /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
                     "
     polyA_signal    3218..3339
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3255..3274)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3309..3328
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3388..3404)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3388..3404)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3401..3423)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3412..3428
                     /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(3436..3442,3443..3460,3461..3466))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3481..3502
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3619..3636)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3790..4378)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3870..3889)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4549..5340,5341..5409))
                     /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     5172..5191
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(5410..5431,1..83))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
       61 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga
      121 tcgggagatc tcccgatccc ctatggtgca ctctcagtac aatctgctct gatgccgcat
      181 agttaagcca gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca
      241 aaatttaagc tacaacaagg caaggcttga ccgacaattg catgaagaat ctgcttaggg
      301 ttaggcgttt tgcgctgctt cgcgatgtac gggccagata tacgcgttga cattgattat
      361 tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt
      421 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
      481 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
      541 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
      601 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc
      661 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
      721 ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac
      781 ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc
      841 aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc
      901 gtgtacggtg ggaggtctat ataagcagag ctctccctat cagtgataga gatctcccta
      961 tcagtgatag agatcgtcga cgagctcgtt tagtgaaccg tcagatcgcc tggagacgcc
     1021 atccacgctg ttttgacctc catagaagac accgggaccg atccagcctc cggactctag
     1081 cgtttaaact taagatgaag ggaaacgaca tcttcgaggc ccagaagatc gagtggcacg
     1141 agggctccgg ctccacctcc ggctccggca tgcagatctt cgtgaagact ctgactggta
     1201 agaccatcac cctcgaggtt gagcccagtg acaccattga gaatgtcaag gcaaagatcc
     1261 aagataagga aggcatccct cctgaccagc agaggctgat ctttgctgga aaacagctgg
     1321 aagatgggcg caccctgtct gactacaaca tccagaaaga gtccaccctg cacctggtac
     1381 tccgtctcag aggtgggtaa taaggatcca ctagtccagt gtggtggaat tctgcagata
     1441 tccagcacag tggcggccgc tcgagtctag agggcccgtt taaacccgct gatcagcctc
     1501 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac
     1561 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg
     1621 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga
     1681 ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga
     1741 aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc
     1801 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     1861 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     1921 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     1981 acttgattag ggtgatggtt cacgtaccta gaagttccta ttccgaagtt cctattctct
     2041 agaaagtata ggaacttcct tggccaaaaa gcctgaactc accgcgacgt ctgtcgagaa
     2101 gtttctgatc gaaaagttcg acagcgtctc cgacctgatg cagctctcgg agggcgaaga
     2161 atctcgtgct ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg taaatagctg
     2221 cgccgatggt ttctacaaag atcgttatgt ttatcggcac tttgcatcgg ccgcgctccc
     2281 gattccggaa gtgcttgaca ttggggaatt cagcgagagc ctgacctatt gcatctcccg
     2341 ccgtgcacag ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg ctgttctgca
     2401 gccggtcgcg gaggccatgg atgcgatcgc tgcggccgat cttagccaga cgagcgggtt
     2461 cggcccattc ggaccgcaag gaatcggtca atacactaca tggcgtgatt tcatatgcgc
     2521 gattgctgat ccccatgtgt atcactggca aactgtgatg gacgacaccg tcagtgcgtc
     2581 cgtcgcgcag gctctcgatg agctgatgct ttgggccgag gactgccccg aagtccggca
     2641 cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg gacaatggcc gcataacagc
     2701 ggtcattgac tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt
     2761 cttctggagg ccgtggttgg cttgtatgga gcagcagacg cgctacttcg agcggaggca
     2821 tccggagctt gcaggatcgc cgcggctccg ggcgtatatg ctccgcattg gtcttgacca
     2881 actctatcag agcttggttg acggcaattt cgatgatgca gcttgggcgc agggtcgatg
     2941 cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt acacaaatcg cccgcagaag
     3001 cgcggccgtc tggaccgatg gctgtgtaga agtactcgcc gatagtggaa accgacgccc
     3061 cagcactcgt ccgagggcaa aggaatagca cgtactacga gatttcgatt ccaccgccgc
     3121 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca
     3181 gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa
     3241 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     3301 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac
     3361 ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
     3421 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     3481 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     3541 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     3601 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     3661 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     3721 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     3781 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     3841 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     3901 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     3961 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     4021 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     4081 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     4141 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     4201 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     4261 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     4321 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     4381 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     4441 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     4501 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     4561 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     4621 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     4681 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     4741 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     4801 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     4861 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     4921 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     4981 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     5041 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     5101 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     5161 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     5221 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     5281 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     5341 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     5401 aatactcata ctcttccttt ttcaatatta t
//
