LOCUS       sequence-429093-        4959 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 4959)
  AUTHORS   Dantuma NP, Lindsten K, Glas R, Jellne M, Masucci MG
  TITLE     Short-lived green fluorescent proteins for quantifying 
            ubiquitin/proteasome-dependent proteolysis in living cells.
  JOURNAL   Nat Biotechnol. 2000 May . 18(5):538-43.
  PUBMED    10802622
REFERENCE   2  (bases 1 to 4959)
  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 
            Biotechnol. 2000 May . 18(5):538-43."
COMMENT     Sequence Label: Ub-R-GFP
FEATURES             Location/Qualifiers
     source          1..4959
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     278..297
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        552..855
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        856..1059
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1010..1030
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             1129..1356
                     /codon_start=1
                     /product="human ubiquitin"
                     /label=ubiquitin
                     /translation="MQIFVKTLTGKTITLEVEPSDTIENVKAKIQDKEGIPPDQQRLIF
                     AGKQLEDGRTLSDYNIQKESTLHLVLRLRGG"
     regulatory      1390..1399
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1396..1398,1399..1401,1402..2115)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1396..1398,1399..1401,1402..2112)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1441..1462)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1702..1721)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2049..2070
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2238..2359
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2275..2294)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2329..2348
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2366..2821)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2503..2524)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2715..2734
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2848..2952
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2954..3311
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2954..3145
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2977..2997)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3162..3297
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3346..4140
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3346..4140
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3400..3419)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4010..4029
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4328..4347)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4372..4419
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      join(4748..4959,1..377)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
       61 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
      121 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
      181 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
      241 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
      301 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
      361 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
      421 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
      481 ccgccatgca ttagttatta atagtaatca attacggggt cattagttca tagcccatat
      541 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
      601 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
      661 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
      721 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
      781 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
      841 atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt
      901 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac
      961 caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc
     1021 ggtaggcgtg tacggtggga ggtctatata agcagagctg gtttagtgaa ccgtcagatc
     1081 cgctagcgct accggactca gatctcgagc tcaagcttcg aattcaccat gcagatcttc
     1141 gtgaagactc tgactggtaa gaccatcacc ctcgaggttg agcccagtga caccattgag
     1201 aatgtcaagg caaagatcca agataaggaa ggcatccctc ctgaccagca gaggctgatc
     1261 tttgctggaa aacagctgga agatgggcgc accctgtctg actacaacat ccagaaagag
     1321 tccaccctgc acctggtact ccgtctcaga ggtgggcgcg ggaagcttgg tcgacaggat
     1381 ccaccggtcg ccaccatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc
     1441 ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag
     1501 ggcgatgcca cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc
     1561 gtgccctggc ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac
     1621 cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag
     1681 gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc
     1741 gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc
     1801 aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc
     1861 gacaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc
     1921 agcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg
     1981 ctgcccgaca accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag
     2041 cgcgatcaca tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac
     2101 gagctgtaca agtaaagcgg ccgcgactct agatcataat cagccatacc acatttgtag
     2161 aggttttact tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga
     2221 atgcaattgt tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     2281 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     2341 aactcatcaa tgtatcttaa ggcgtaaatt gtaagcgtta atattttgtt aaaattcgcg
     2401 ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct
     2461 tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt
     2521 ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat
     2581 ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca
     2641 ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac
     2701 gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta
     2761 gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg
     2821 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     2881 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     2941 aaaaggaaga gtcctgaggc ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg
     3001 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     3061 caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     3121 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     3181 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     3241 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     3301 gcttttgcaa agatcgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat
     3361 ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca
     3421 caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg
     3481 gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcaaga cgaggcagcg
     3541 cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact
     3601 gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct
     3661 caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg
     3721 cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt
     3781 actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc
     3841 gcgccagccg aactgttcgc caggctcaag gcgagcatgc ccgacggcga ggatctcgtc
     3901 gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga
     3961 ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc
     4021 cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt
     4081 atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga
     4141 gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt
     4201 tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg
     4261 gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac cctaggggga
     4321 ggctaactga aacacggaag gagacaatac cggaaggaac ccgcgctatg acggcaataa
     4381 aaagacagaa taaaacgcac ggtgttgggt cgtttgttca taaacgcggg gttcggtccc
     4441 agggctggca ctctgtcgat accccaccga gaccccattg gggccaatac gcccgcgttt
     4501 cttccttttc cccaccccac cccccaagtt cgggtgaagg cccagggctc gcagccaacg
     4561 tcggggcggc aggccctgcc atagcctcag gttactcata tatactttag attgatttaa
     4621 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     4681 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     4741 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     4801 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     4861 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc
     4921 accacttcaa gaactctgta gcaccgccta catacctcg
//
