LOCUS       sequence-190680-        7528 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 7528)
  AUTHORS   Matsunaga K, Saitoh T, Tabata K, Omori H, Satoh T, Kurotori N, 
            Maejima I, Shirahama-Noda K, Ichimura T, Isobe T, Akira S, Noda T, 
            Yoshimori T
  TITLE     Two Beclin 1-binding proteins, Atg14L and Rubicon, reciprocally 
            regulate autophagy at different stages.
  JOURNAL   Nat Cell Biol. 2009 Apr . 11(4):385-96.
  PUBMED    19270696
REFERENCE   2  (bases 1 to 7528)
  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 Cell 
            Biol. 2009 Apr . 11(4):385-96."
COMMENT     Sequence Label: pEGFP-Rubicon
FEATURES             Location/Qualifiers
     source          1..7528
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        100..291
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     248..268
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        complement(293..397)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      424..879
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(511..530)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     721..742
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    886..1007
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(897..916)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     951..970
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(join(3994..4704,4705..4707,4708..4710))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(3994..4704,4705..4707,4708..4710))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4036..4057)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     4385..4404
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4644..4665
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      4707..4716
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(4755..4958)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(4784..4804)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        4959..5262
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     rep_origin      complement(5437..6025)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5517..5536)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    6354..6401
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     6426..6445
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(6633..7427)
                     /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             complement(6633..7427)
                     /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(6744..6763)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     7354..7373
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(join(7462..7528,1..291))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(7476..7528,1..83)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 gcctcggcct ctgcataaat aaaaaaaatt agtcagccat ggggcggaga atgggcggaa
       61 ctgggcggag ttaggggcgg gatgggcgga gttaggggcg ggactatggt tgctgactaa
      121 ttgagatgca tgctttgcat acttctgcct gctggggagc ctggggactt tccacacctg
      181 gttgctgact aattgagatg catgctttgc atacttctgc ctgctgggga gcctggggac
      241 tttccacacc ctaactgaca cacattccac agctggttct ttccgcctca ggactcttcc
      301 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
      361 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
      421 ctgacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
      481 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
      541 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
      601 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
      661 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
      721 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
      781 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
      841 ttaacgcgaa ttttaacaaa atattaacgc ttacaattta cgcgttaaga tacattgatg
      901 agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg
      961 atgctattgc tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt
     1021 gcattcattt tatgtttcag gttcaggggg aggtgtggga ggttttttaa agcaagtaaa
     1081 acctctacaa atgtggtatg gctgattatg atcagttatc tagatccggt ggatcccatc
     1141 tcaggtggtc tctaggacgg tggcctctag agccaaggct tctgtgggct cctcctcata
     1201 gtcagacagg taggactcca ggctctgctt ggccagcagc tcccgtcggg cctgcagccg
     1261 ctcacaccgg gggcagcgtc cagacttgaa gcaagtttta tggtaacacg ctttacactc
     1321 ttcacaagtc cggcacttat gcagctcaaa aggaaagatg acatcctcct cattctgaca
     1381 gaattcacag atgaagccct tggcctgaca cagcatgcat ctctccacgt gggcggctcc
     1441 cgctctagtg agctccgcca gccggggccc cagttctccc ttcttggttg cagtcaggtc
     1501 actcagcgag tacagatgga ggtcctctgt caggtggcct ggaaccacgt caaaggaatc
     1561 caggagctct ttggccagtc ggcatgtctt aaacatgttc ttcatgtggt acagctgaac
     1621 ccgcagcagc cggacttggt taagcagctt gaccttcctg tagagtgcgc tattgatgtc
     1681 ctgcacattg aagagaggat cattccagat cttcaggagc aggtccttag agaagttgct
     1741 gacatagtac ttgctgaagt cccatttgcg cagaatgcgg ctggggacga ccatctgggc
     1801 gttctcgtgg cagcactggc agaagtactt gcctaggtac tcacagtacc gcagccgctt
     1861 gatatagtcg gggtccgtcc ggatgccaca tcccgcacag cggtaattct gcttggccac
     1921 ggcaatcttc ctcgtggggg ctggatgaac attaaaaatg atctgcggcc ggggcggagc
     1981 ccactccagg ttgccacgga cgcggattcg cagcttgtag atgtccgcgt gctgcccgtc
     2041 atctggtgag atgggcaggg agtcggggat gggtaggagc ttctgtggag catcatgctc
     2101 tgggactagc cactcgggct ccgaggcagc tggaagctgc atcccctcaa actgcttcag
     2161 aagccccatg gccactgcct cagccgatgt ggagtgcagg aagcagtgcg agagattctg
     2221 ggagaaggat gatttgccgt ttgagactgc actcctccgg atgtcagcgt cttggatttc
     2281 caagtcgtca gcatcctcag cagagccgga ctcagagagc tgcgtcgaat cccgtgaact
     2341 gaactgggag ctgctggagg tgacccggaa gcttccattc tcagtctccc ggtacgcagg
     2401 gagcaggttt ttggtgcgga tctgctggcg ccgaaggcgg atcttctgct tcagttcctg
     2461 gatctctcgg tcactatctt cctcctctac ttcctcctct tctagacact ggctcatcat
     2521 gttgcacttc atcagctcaa tggcggcaat caaggactcg gagatgctga agtgggcgtt
     2581 ttccttttcc aggtctgcac agctgccaaa gtcttgctca gagaggtaac tgatgaggga
     2641 ctgtccttct gatggtcttc taaacatgcc ttctcctgag cagaggtact gaccaccctc
     2701 tgcggctcct ctggaggcaa tgttggtgtc cgagtgggag cgaatatggc tcttctttgt
     2761 cccgctggcg acaggggctg tctgcccttc tgagaagctg gacctgcgaa ggacgctgcc
     2821 tgcctgactc tgcctgcctt cctcttggtc ccctagagaa gaggcagcag actccagctt
     2881 ttgagagctg ctggaggaga acaagtgaga gctgctgctc tcgccgttgc tgctgtgact
     2941 ctcacaactg gcagtccggc catgaggggc tgggttgcca atggccaggt actcgggccc
     3001 ctcactgaca tcacttgggg agtcattctg gctgctgacc caggatgcct ctagggggct
     3061 agtcagagtg ctgtcgctca tctcctgtgc tgtcagtggg gaatcctggg ctagagtaga
     3121 gtctgaaggc agcggagctg gggtggtttg agtctctgct ggtgagaggc attctctgtc
     3181 ttcttgaggt tgccaggaag ggccagagag tgaaaaggaa gtagatcttc tctctgagct
     3241 gtggctggac tgtggcatag actgaaggct ggaggaggac ccaaagtagg aatgctgagc
     3301 atagctagct ggaggggtgt atgtggaacc aggcagagcg gtcagactct ggctctttgt
     3361 gaccagcagc gggctctcct gctttctggc aaacatggat gcatcgatct gagccagaag
     3421 acgggggttg ttctgttcca ctgcttccag gcactggagc atggctgtga cgtgggcgtc
     3481 actcagcagg aaggcagttt ctgtgtagaa ttttctgatg tactgtctgt ccccgagcag
     3541 aggccggagc tgggctgaga ggcagtggca ctgcaggcta tgctgcagcc acagctctgc
     3601 aacagcacgc tcactcacgc tgtcagtgtc gctctggtcg ctctcatgca aactgatgaa
     3661 cttctccacg tgaagggctg agtgcgggct gagccaccga atgtctttca caaactgcca
     3721 gtaatcagcc tggcggcaac acacctggtc atggatgagc ccatgataca ggatgttctg
     3781 catgtccctg cacagccgct ccaggccacc atacttggac cagacattgg ggcagttggc
     3841 tgacaccaaa ccttccacag tcgtcttcaa attacccagc agctgccagt gctccctcct
     3901 gctcttctct agcagacgct ccccatcgcc gccgccaaga tccattcccg cgccctccgg
     3961 acgcatggat ctcgagatct gagtccggcc ggacttgtac agctcgtcca tgccgagagt
     4021 gatcccggcg gcggtcacga actccagcag gaccatgtga tcgcgcttct cgttggggtc
     4081 tttgctcagg gcggactggg tgctcaggta gtggttgtcg ggcagcagca cggggccgtc
     4141 gccgatgggg gtgttctgct ggtagtggtc ggcgagctgc acgctgccgt cctcgatgtt
     4201 gtggcggatc ttgaagttca ccttgatgcc gttcttctgc ttgtcggcca tgatatagac
     4261 gttgtggctg ttgtagttgt actccagctt gtgccccagg atgttgccgt cctccttgaa
     4321 gtcgatgccc ttcagctcga tgcggttcac cagggtgtcg ccctcgaact tcacctcggc
     4381 gcgggtcttg tagttgccgt cgtccttgaa gaagatggtg cgctcctgga cgtagccttc
     4441 gggcatggcg gacttgaaga agtcgtgctg cttcatgtgg tcggggtagc ggctgaagca
     4501 ctgcacgccg taggtcaggg tggtcacgag ggtgggccag ggcacgggca gcttgccggt
     4561 ggtgcagatg aacttcaggg tcagcttgcc gtaggtggca tcgccctcgc cctcgccgga
     4621 cacgctgaac ttgtggccgt ttacgtcgcc gtccagctcg accaggatgg gcaccacccc
     4681 ggtgaacagc tcctcgccct tgctcaccat ggtggcgacc ggtagcgcta gcggatctga
     4741 cggttcacta aaccagctct gcttatatag acctcccacc gtacacgcct accgcccatt
     4801 tgcgtcaatg gggcggagtt gttacgacat tttggaaagt cccgttgatt ttggtgccaa
     4861 aacaaactcc cattgacgtc aatggggtgg agacttggaa atccccgtga gtcaaaccgc
     4921 tatccacgcc cattgatgta ctgccaaaac cgcatcacca tggtaatagc gatgactaat
     4981 acgtagatgt actgccaagt aggaaagtcc cataaggtca tgtactgggc ataatgccag
     5041 gcgggccatt taccgtcatt gacgtcaata gggggcgtac ttggcatatg atacacttga
     5101 tgtactgcca agtgggcagt ttaccgtaaa tactccaccc attgacgtca atggaaagtc
     5161 cctattggcg ttactatggg aacatacgtc attattgacg tcaatgggcg ggggtcgttg
     5221 ggcggtcagc caggcgggcc atttaccgta agttatgtaa cgcggaactc catatatggg
     5281 ctatgaacta atgaccccgt aattgattac tattaataac taatgcatgg cggtaatacg
     5341 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     5401 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     5461 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     5521 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     5581 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     5641 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     5701 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     5761 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     5821 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     5881 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     5941 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     6001 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     6061 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     6121 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     6181 acctgaggct atggcagggc ctgccgcccc gacgttggct gcgagccctg ggccttcacc
     6241 cgaacttggg gggtggggtg gggaaaagga agaaacgcgg gcgtattggc cccaatgggg
     6301 tctcggtggg gtatcgacag agtgccagcc ctgggaccga accccgcgtt tatgaacaaa
     6361 cgacccaaca ccgtgcgttt tattctgtct ttttattgcc gtcatagcgc gggttccttc
     6421 cggtattgtc tccttccgtg tttcagttag cctcccccta gggtgggcga agaactccag
     6481 catgagatcc ccgcgctgga ggatcatcca gccggcgtcc cggaaaacga ttccgaagcc
     6541 caacctttca tagaaggcgg cggtggaatc gaaatctcgt gatggcaggt tgggcgtcgc
     6601 ttggtcggtc atttcgaacc ccagagtccc gctcagaaga actcgtcaag aaggcgatag
     6661 aaggcgatgc gctgcgaatc gggagcggcg ataccgtaaa gcacgaggaa gcggtcagcc
     6721 cattcgccgc caagctcttc agcaatatca cgggtagcca acgctatgtc ctgatagcgg
     6781 tccgccacac ccagccggcc acagtcgatg aatccagaaa agcggccatt ttccaccatg
     6841 atattcggca agcaggcatc gccatgggtc acgacgagat cctcgccgtc gggcatgctc
     6901 gccttgagcc tggcgaacag ttcggctggc gcgagcccct gatgctcttc gtccagatca
     6961 tcctgatcga caagaccggc ttccatccga gtacgtgctc gctcgatgcg atgtttcgct
     7021 tggtggtcga atgggcaggt agccggatca agcgtatgca gccgccgcat tgcatcagcc
     7081 atgatggata ctttctcggc aggagcaagg tgagatgaca ggagatcctg ccccggcact
     7141 tcgcccaata gcagccagtc ccttcccgct tcagtgacaa cgtcgagcac agctgcgcaa
     7201 ggaacgcccg tcgtggccag ccacgatagc cgcgctgcct cgtcttgcag ttcattcagg
     7261 gcaccggaca ggtcggtctt gacaaaaaga accgggcgcc cctgcgctga cagccggaac
     7321 acggcggcat cagagcagcc gattgtctgt tgtgcccagt catagccgaa tagcctctcc
     7381 acccaagcgg ccggagaacc tgcgtgcaat ccatcttgtt caatcatgcg aaacgatcct
     7441 catcctgtct cttgatcgat ctttgcaaaa gcctaggcct ccaaaaaagc ctcctcacta
     7501 cttctggaat agctcagagg ccgaggcg
//
