LOCUS       sequence-43564-e        7883 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7883)
  AUTHORS   Sims JJ, Scavone F, Cooper EM, Kane LA, Youle RJ, Boeke JD, Cohen RE
  TITLE     Polyubiquitin-sensor proteins reveal localization and linkage-type 
            dependence of cellular ubiquitin signaling.
  JOURNAL   Nat Methods. 2012 Feb 5. doi: 10.1038/nmeth.1888.
  PUBMED    22306808
REFERENCE   2  (bases 1 to 7883)
  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: "Nat 
            Methods. 2012 Feb 5. doi: 10.1038/nmeth.1888."
COMMENT     Sequence Label: pQCXIP-Vx3K0-mEGFP
FEATURES             Location/Qualifiers
     source          1..7883
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        310..513
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             514..689
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    752..951
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1152..1568
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1498..1520
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1536..1552
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     enhancer        1605..1908
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1909..2112
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2109..2133
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      2254..2263
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2272..2298
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             2272..2298
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     regulatory      2554..2563
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2560..2562,2563..2565,2566..3279)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKVSKDPNEKRDHMVLL
                     EFVTAAGITLGXDELYK"
     primer_bind     complement(2605..2626)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2866..2885)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3213..3234
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    3288..3860
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3455..3472)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3682..3701
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3875..4474
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3875..3894)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4371..4391
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4589..5014
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     primer_bind     complement(5117..5139)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5275..5295)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5280..5609
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5460..5595
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5631..5650
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5731..5748)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5902..6490)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5982..6001)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6661..7452,7453..7521))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     7284..7303
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7522..7626)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7694..7712
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     enhancer        join(7812..7883,1..309)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
       61 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
      121 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
      181 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgctctg gcattatgcc
      241 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
      301 attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca
      361 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
      421 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg
      481 cgtgtacggt gggaggtcta tataagcaga gctcaataaa agagcccaca acccctcact
      541 cggcgcgcca gtcttccgat agactgcgtc gcccgggtac ccgtattccc aataaagcct
      601 cttgctgttt gcatccgaat cgtggtctcg ctgttccttg ggagggtctc ctctgagtga
      661 ttgactaccc acgacggggg tctttcattt gggggctcgt ccgggatttg gagacccctg
      721 cccagggacc accgacccac caccgggagg taagctggcc agcaacttat ctgtgtctgt
      781 ccgattgtct agtgtctatg tttgatgtta tgcgcctgcg tctgtactag ttagctaact
      841 agctctgtat ctggcggacc cgtggtggaa ctgacgagtt ctgaacaccc ggccgcaacc
      901 ctgggagacg tcccagggac tttgggggcc gtttttgtgg cccgacctga ggaagggagt
      961 cgatgtggaa tccgaccccg tcaggatatg tggttctggt aggagacgag aacctaaaac
     1021 agttcccgcc tccgtctgaa tttttgcttt cggtttggaa ccgaagccgc gcgtcttgtc
     1081 tgctgcagcg ctgcagcatc gttctgtgtt gtctctgtct gactgtgttt ctgtatttgt
     1141 ctgaaaatta gggccagact gttaccactc ccttaagttt gaccttaggt cactggaaag
     1201 atgtcgagcg gatcgctcac aaccagtcgg tagatgtcaa gaagagacgt tgggttacct
     1261 tctgctctgc agaatggcca acctttaacg tcggatggcc gcgagacggc acctttaacc
     1321 gagacctcat cacccaggtt aagatcaagg tcttttcacc tggcccgcat ggacacccag
     1381 accaggtccc ctacatcgtg acctgggaag ccttggcttt tgacccccct ccctgggtca
     1441 agccctttgt acaccctaag cctccgcctc ctcttcctcc atccgccccg tctctccccc
     1501 ttgaacctcc tcgttcgacc ccgcctcgat cctcccttta tccagccctc actccttctc
     1561 taggcgccgg aattgaagat ctgggggatc gatcctctag agtccgttac ataacttacg
     1621 gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg
     1681 tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta
     1741 cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt
     1801 gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac
     1861 tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt
     1921 tggcagtaca tcaatgggcg tgaatagcgg tttgactcac ggggatttcc aagtctccac
     1981 cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt
     2041 cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat
     2101 ataagcagag ctcgtttagt gaaccgtcag atcgcctgga gacgccatcc acgctgtttt
     2161 gacctccata gaagacaccg ggaccgatcc agcctccgcg gccgggaacg gtgcattgga
     2221 acggacctgc aggaattgat ccgcggccgc gccgccacca tggagggcag ctacccctac
     2281 gacgtgcccg actacgccgg cggcagcggc ctcgagatgg aagaagaaga agatcctgat
     2341 ttacgcgctg ctatccagga atctctgcgt gaagctcgtg ctgaagaacg cgttcgcgaa
     2401 gacgaagaag aactgatccg tcgcgctatc gaactgtctc tgcgcgaatc tcgtgaagtt
     2461 aacgctcagg aagaggacga ggaggagctg atccgccgcg ccatcgagct gagcctgcgc
     2521 gagagccgta actctgctgg ggatccaccg gtcgccacca tggtgagcaa gggcgaggag
     2581 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
     2641 ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc
     2701 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac
     2761 ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
     2821 gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac
     2881 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
     2941 ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac
     3001 agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag
     3061 atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc
     3121 cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccaag
     3181 gtgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc
     3241 gccgggatca ctctcggcan ggacgagctg tacaagtaag aattccgccc ctctccctcc
     3301 ccccccccta acgttactgg ccgaagccgc ttggaataag gccggtgtgc gtttgtctat
     3361 atgttatttt ccaccatatt gccgtctttt ggcaatgtga gggcccggaa acctggccct
     3421 gtcttcttga cgagcattcc taggggtctt tcccctctcg ccaaaggaat gcaaggtctg
     3481 ttgaatgtcg tgaaggaagc agttcctctg gaagcttctt gaagacaaac aacgtctgta
     3541 gcgacccttt gcaggcagcg gaacccccca cctggcgaca ggtgcctctg cggccaaaag
     3601 ccacgtgtat aagatacacc tgcaaaggcg gcacaacccc agtgccacgt tgtgagttgg
     3661 atagttgtgg aaagagtcaa atggctctcc tcaagcgtat tcaacaaggg gctgaaggat
     3721 gcccagaagg taccccattg tatgggatct gatctggggc ctcggtgcac atgctttaca
     3781 tgtgtttagt cgaggttaaa aaacgtctag gccccccgaa ccacggggac gtggttttcc
     3841 tttgaaaaac acgatgataa gcttgatcac gcgtatgacc gagtacaagc ccacggtgcg
     3901 cctcgccacc cgcgacgacg tccccagggc cgtacgcacc ctcgccgccg cgttcgccga
     3961 ctaccccgcc acgcgccaca ccgtcgatcc ggaccgccac atcgagcggg tcaccgagct
     4021 gcaagaactc ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga
     4081 cggcgccgcg gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc
     4141 cgagatcggc ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc agcaacagat
     4201 ggaaggcctc ctggcgccgc accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg
     4261 cgtctcgccc gaccaccagg gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga
     4321 ggcggccgag cgcgccgggg tgcccgcctt cctggagacc tccgcgcccc gcaacctccc
     4381 cttctacgag cggctcggct tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg
     4441 cacctggtgc atgacccgca agcccggtgc ctgagatatc agtggtccag gctctagttt
     4501 tgactcaaca atatcaccag ctgaagccta tagagtacga gccatagata aaataaaaga
     4561 ttttatttag tctccagaaa aaggggggaa tgaaagaccc cacctgtagg tttggcaagc
     4621 tagcttaagt aacgccattt tgcaaggcat ggaaaaatac ataactgaga atagagaagt
     4681 tcagatcaag gtcaggaaca gatggaacag ggtcgaccct agagaaccat cagatgtttc
     4741 cagggtgccc caaggacctg aaatgaccct gtgccttatt tgaactaacc aatcagttcg
     4801 cttctcgctt ctgttcgcgc gcttctgctc cccgagctca ataaaagagc ccacaacccc
     4861 tcactcgggg cgccagtcct ccgattgact gagtcgcccg ggtacccgtg tatccaataa
     4921 accctcttgc agttgcatcc gacttgtggt ctcgctgttc cttgggaggg tctcctctga
     4981 gtgattgact acccgtcagc gggggtcttt catttggggg ctcgtccggg atcgggagac
     5041 ccctgcccag ggaccaccga cccaccaccg ggaggtaagc tggctgcctc gcgcgtttcg
     5101 gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt
     5161 aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc
     5221 ggggcgcagc catgacccag tcacgtagcg atagcggagt gtagatccgg ctgtggaatg
     5281 tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     5341 tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa
     5401 gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta actccgccca
     5461 tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt
     5521 ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag
     5581 gcttttttgg aggcctaggc ttttgcaaaa agcttactgg cttaactatg cggcatcaga
     5641 gcagattgta ctgagagtgc accatatgcg gtgtgaaata ccgcacagat gcgtaaggag
     5701 aaaataccgc atcaggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     5761 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     5821 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     5881 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     5941 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     6001 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     6061 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     6121 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     6181 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     6241 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     6301 agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat ttggtatctg
     6361 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     6421 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     6481 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     6541 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     6601 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     6661 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     6721 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     6781 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     6841 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     6901 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     6961 cgttgttgcc attgctgcag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     7021 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     7081 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     7141 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     7201 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     7261 ctcttgcccg gcgtcaacac gggataatac cgcgccacat agcagaactt taaaagtgct
     7321 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     7381 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     7441 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     7501 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     7561 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     7621 tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac
     7681 attaacctat aaaaataggc gtatcacgag gccctttcgt cttcaagaat tagcttggcc
     7741 attgcatacg ttgtatccat atcataatat gtacatttat attggctcat gtccaacatt
     7801 accgccatgt tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt
     7861 agttcatagc ccatatatgg agt
//
