LOCUS       sequence-43567-e        7919 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 7919)
  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 7919)
  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-NLS-Vx3-mEGFP
FEATURES             Location/Qualifiers
     source          1..7919
                     /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"
     CDS             2266..2286
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2311..2331
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     regulatory      2590..2599
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2596..2598,2599..2601,2602..3315)
                     /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(2641..2662)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2902..2921)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3249..3270
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    3324..3896
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3491..3508)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3718..3737
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3911..4510
                     /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(3911..3930)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4407..4427
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4625..5050
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     primer_bind     complement(5153..5175)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5311..5331)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5316..5645
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5496..5631
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5667..5686
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5767..5784)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5938..6526)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6018..6037)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6697..7488,7489..7557))
                     /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     7320..7339
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7558..7662)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7730..7748
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     enhancer        join(7848..7919,1..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 tgagccccaa gaagaagcgc
     2281 aaggtggagg ccagcaagat gaccgcctct ccgaaaaaaa aacgtaaagt tgaagcttct
     2341 aaaatgaccg ctatggaaga agaagaagat cctgatttaa aagctgctat ccaggaatct
     2401 ctgcgtgaag ctcgtgctga agaaaaagtt aaagaagacg aagaagaact gatccgtaaa
     2461 gctatcgaac tgtctctgaa agaatctcgt gaagttaacg ctcaggaaga ggacgaggag
     2521 gagctgatcc gcaaggccat cgagctgagc ctgaaggagt cccgtaactc tgctggggat
     2581 ccaccggtcg ccaccatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc
     2641 ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag
     2701 ggcgatgcca cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc
     2761 gtgccctggc ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac
     2821 cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag
     2881 gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc
     2941 gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc
     3001 aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc
     3061 gacaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc
     3121 agcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg
     3181 ctgcccgaca accactacct gagcacccag tccaaggtga gcaaagaccc caacgagaag
     3241 cgcgatcaca tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcanggac
     3301 gagctgtaca agtaagaatt ccgcccctct ccctcccccc cccctaacgt tactggccga
     3361 agccgcttgg aataaggccg gtgtgcgttt gtctatatgt tattttccac catattgccg
     3421 tcttttggca atgtgagggc ccggaaacct ggccctgtct tcttgacgag cattcctagg
     3481 ggtctttccc ctctcgccaa aggaatgcaa ggtctgttga atgtcgtgaa ggaagcagtt
     3541 cctctggaag cttcttgaag acaaacaacg tctgtagcga ccctttgcag gcagcggaac
     3601 cccccacctg gcgacaggtg cctctgcggc caaaagccac gtgtataaga tacacctgca
     3661 aaggcggcac aaccccagtg ccacgttgtg agttggatag ttgtggaaag agtcaaatgg
     3721 ctctcctcaa gcgtattcaa caaggggctg aaggatgccc agaaggtacc ccattgtatg
     3781 ggatctgatc tggggcctcg gtgcacatgc tttacatgtg tttagtcgag gttaaaaaac
     3841 gtctaggccc cccgaaccac ggggacgtgg ttttcctttg aaaaacacga tgataagctt
     3901 gatcacgcgt atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc
     3961 cagggccgta cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt
     4021 cgatccggac cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt
     4081 cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac
     4141 cacgccggag agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga
     4201 gttgagcggt tcccggctgg ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg
     4261 gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa
     4321 gggtctgggc agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc
     4381 cgccttcctg gagacctccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac
     4441 cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc
     4501 cggtgcctga gatatcagtg gtccaggctc tagttttgac tcaacaatat caccagctga
     4561 agcctataga gtacgagcca tagataaaat aaaagatttt atttagtctc cagaaaaagg
     4621 ggggaatgaa agaccccacc tgtaggtttg gcaagctagc ttaagtaacg ccattttgca
     4681 aggcatggaa aaatacataa ctgagaatag agaagttcag atcaaggtca ggaacagatg
     4741 gaacagggtc gaccctagag aaccatcaga tgtttccagg gtgccccaag gacctgaaat
     4801 gaccctgtgc cttatttgaa ctaaccaatc agttcgcttc tcgcttctgt tcgcgcgctt
     4861 ctgctccccg agctcaataa aagagcccac aacccctcac tcggggcgcc agtcctccga
     4921 ttgactgagt cgcccgggta cccgtgtatc caataaaccc tcttgcagtt gcatccgact
     4981 tgtggtctcg ctgttccttg ggagggtctc ctctgagtga ttgactaccc gtcagcgggg
     5041 gtctttcatt tgggggctcg tccgggatcg ggagacccct gcccagggac caccgaccca
     5101 ccaccgggag gtaagctggc tgcctcgcgc gtttcggtga tgacggtgaa aacctctgac
     5161 acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag
     5221 cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg acccagtcac
     5281 gtagcgatag cggagtgtag atccggctgt ggaatgtgtg tcagttaggg tgtggaaagt
     5341 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     5401 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     5461 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     5521 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     5581 cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     5641 gcaaaaagct tactggctta actatgcggc atcagagcag attgtactga gagtgcacca
     5701 tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgctcttc
     5761 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     5821 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     5881 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     5941 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     6001 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     6061 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     6121 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     6181 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     6241 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     6301 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     6361 ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     6421 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     6481 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     6541 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     6601 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     6661 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     6721 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     6781 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     6841 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     6901 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     6961 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctgcaggcat
     7021 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     7081 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     7141 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     7201 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     7261 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caacacggga
     7321 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     7381 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     7441 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     7501 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     7561 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     7621 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     7681 gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa ataggcgtat
     7741 cacgaggccc tttcgtcttc aagaattagc ttggccattg catacgttgt atccatatca
     7801 taatatgtac atttatattg gctcatgtcc aacattaccg ccatgttgac attgattatt
     7861 gactagttat taatagtaat caattacggg gtcattagtt catagcccat atatggagt
//
