LOCUS       sequence-43452-e        7832 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 7832)
  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 7832)
  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-Rx3(A7)-mEGFP
FEATURES             Location/Qualifiers
     source          1..7832
                     /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"
     regulatory      2503..2512
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2509..2511,2512..2514,2515..3228)
                     /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(2554..2575)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2815..2834)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3162..3183
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    3237..3809
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3404..3421)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3631..3650
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3824..4423
                     /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(3824..3843)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4320..4340
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4538..4963
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     primer_bind     complement(5066..5088)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5224..5244)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5229..5558
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5409..5544
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5580..5599
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5680..5697)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5851..6439)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5931..5950)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6610..7401,7402..7470))
                     /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     7233..7252
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7471..7575)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7643..7661
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     enhancer        join(7761..7832,1..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 ccatatgacc
     2281 gaggaggagc aattcgcact cgcactgaag atgagcgagc aagaggcggc ggctgccgct
     2341 gctgccgcag aagaggaaga agaacagttt gctctggctc tcaaaatgag tgaacaggaa
     2401 gctgctgccg cagcggcggc tgccgaggag gaagaagagc tcttgaggaa agccattgca
     2461 gaaagtctga atagttgccg gccttctggg gatccaccgg tcgccaccat ggtgagcaag
     2521 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
     2581 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
     2641 ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
     2701 ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc
     2761 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac
     2821 ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc
     2881 gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac
     2941 aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg
     3001 aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag
     3061 cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc
     3121 cagtccaagg tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc
     3181 gtgaccgccg ccgggatcac tctcggcang gacgagctgt acaagtaaga attccgcccc
     3241 tctccctccc ccccccctaa cgttactggc cgaagccgct tggaataagg ccggtgtgcg
     3301 tttgtctata tgttattttc caccatattg ccgtcttttg gcaatgtgag ggcccggaaa
     3361 cctggccctg tcttcttgac gagcattcct aggggtcttt cccctctcgc caaaggaatg
     3421 caaggtctgt tgaatgtcgt gaaggaagca gttcctctgg aagcttcttg aagacaaaca
     3481 acgtctgtag cgaccctttg caggcagcgg aaccccccac ctggcgacag gtgcctctgc
     3541 ggccaaaagc cacgtgtata agatacacct gcaaaggcgg cacaacccca gtgccacgtt
     3601 gtgagttgga tagttgtgga aagagtcaaa tggctctcct caagcgtatt caacaagggg
     3661 ctgaaggatg cccagaaggt accccattgt atgggatctg atctggggcc tcggtgcaca
     3721 tgctttacat gtgtttagtc gaggttaaaa aacgtctagg ccccccgaac cacggggacg
     3781 tggttttcct ttgaaaaaca cgatgataag cttgatcacg cgtatgaccg agtacaagcc
     3841 cacggtgcgc ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc
     3901 gttcgccgac taccccgcca cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt
     3961 caccgagctg caagaactct tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt
     4021 cgcggacgac ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg aagcgggggc
     4081 ggtgttcgcc gagatcggcc cgcgcatggc cgagttgagc ggttcccggc tggccgcgca
     4141 gcaacagatg gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt ggttcctggc
     4201 caccgtcggc gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc
     4261 cggagtggag gcggccgagc gcgccggggt gcccgccttc ctggagacct ccgcgccccg
     4321 caacctcccc ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga
     4381 aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc tgagatatca gtggtccagg
     4441 ctctagtttt gactcaacaa tatcaccagc tgaagcctat agagtacgag ccatagataa
     4501 aataaaagat tttatttagt ctccagaaaa aggggggaat gaaagacccc acctgtaggt
     4561 ttggcaagct agcttaagta acgccatttt gcaaggcatg gaaaaataca taactgagaa
     4621 tagagaagtt cagatcaagg tcaggaacag atggaacagg gtcgacccta gagaaccatc
     4681 agatgtttcc agggtgcccc aaggacctga aatgaccctg tgccttattt gaactaacca
     4741 atcagttcgc ttctcgcttc tgttcgcgcg cttctgctcc ccgagctcaa taaaagagcc
     4801 cacaacccct cactcggggc gccagtcctc cgattgactg agtcgcccgg gtacccgtgt
     4861 atccaataaa ccctcttgca gttgcatccg acttgtggtc tcgctgttcc ttgggagggt
     4921 ctcctctgag tgattgacta cccgtcagcg ggggtctttc atttgggggc tcgtccggga
     4981 tcgggagacc cctgcccagg gaccaccgac ccaccaccgg gaggtaagct ggctgcctcg
     5041 cgcgtttcgg tgatgacggt gaaaacctct gacacatgca gctcccggag acggtcacag
     5101 cttgtctgta agcggatgcc gggagcagac aagcccgtca gggcgcgtca gcgggtgttg
     5161 gcgggtgtcg gggcgcagcc atgacccagt cacgtagcga tagcggagtg tagatccggc
     5221 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     5281 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     5341 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     5401 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     5461 taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt
     5521 agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gcttactggc ttaactatgc
     5581 ggcatcagag cagattgtac tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg
     5641 cgtaaggaga aaataccgca tcaggcgctc ttccgcttcc tcgctcactg actcgctgcg
     5701 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     5761 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     5821 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     5881 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     5941 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     6001 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     6061 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     6121 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     6181 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     6241 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt
     6301 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     6361 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     6421 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     6481 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     6541 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     6601 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     6661 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     6721 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     6781 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     6841 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     6901 tttgcgcaac gttgttgcca ttgctgcagg catcgtggtg tcacgctcgt cgtttggtat
     6961 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     7021 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     7081 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     7141 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     7201 accgagttgc tcttgcccgg cgtcaacacg ggataatacc gcgccacata gcagaacttt
     7261 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     7321 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     7381 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     7441 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     7501 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     7561 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtctaag aaaccattat
     7621 tatcatgaca ttaacctata aaaataggcg tatcacgagg ccctttcgtc ttcaagaatt
     7681 agcttggcca ttgcatacgt tgtatccata tcataatatg tacatttata ttggctcatg
     7741 tccaacatta ccgccatgtt gacattgatt attgactagt tattaatagt aatcaattac
     7801 ggggtcatta gttcatagcc catatatgga gt
//
