LOCUS       sequence-142595-        6456 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 6456)
  AUTHORS   Chen H, Fre S, Slepnev VI, Capua MR, Takei K, Butler MH, Di Fiore 
            PP, De Camilli P
  TITLE     Epsin is an EH-domain-binding protein implicated in 
            clathrin-mediated endocytosis.
  JOURNAL   Nature. 1998 Aug 20. 394(6695):793-7.
  PUBMED    9723620
REFERENCE   2  (bases 1 to 6456)
  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: "Nature. 
            1998 Aug 20. 394(6695):793-7."
COMMENT     Sequence Label: pEGFPC1-EPSIN 1
FEATURES             Location/Qualifiers
     source          1..6456
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        68..371
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        372..575
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     526..546
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      614..623
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(620..622,623..625,626..1336)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(620..622,623..625,626..1336)
                     /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(665..686)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(926..945)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1273..1294
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3251..3372
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3288..3307)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3342..3361
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3379..3834)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3516..3537)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3728..3747
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3861..3965
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3967..4324
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3967..4158
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3990..4010)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4175..4310
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4359..5153
                     /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             4359..5153
                     /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(4413..4432)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5023..5042
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5341..5360)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5385..5432
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5761..6349
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6250..6269
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 catgcattag ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg
       61 agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc
      121 gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt
      181 gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc
      241 atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg
      301 cccagtacat gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg
      361 ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact
      421 cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa
      481 atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta
      541 ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt cagatccgct
      601 agcgctaccg gtcgccacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc
      661 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg
      721 cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct
      781 gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg
      841 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt
      901 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa
      961 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga
     1021 cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg tctatatcat
     1081 ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga
     1141 cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt
     1201 gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga
     1261 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat
     1321 ggacgagctg tacaagtccg gactcagatc tcgagctcaa gcttcgaatt ctatgtccac
     1381 atcatcgctg cggcgtcaga tgaagaatat cgtccacaac tactcagagg cagagatcaa
     1441 ggtccgggag gccacaagca atgatccctg gggcccatcc agctccctca tgtcagagat
     1501 tgctgacctc acctacaatg ttgtggcctt ctccgagatt atgagcatga tctggaagcg
     1561 gctcaacgac catggcaaga attggaggca cgtctacaag gccatgacgt tgatggagta
     1621 cctcatcaag actggctctg agcgtgtatc acaacagtgc aaagagaaca tgtatgctgt
     1681 gcagacgctg aaggacttcc aatatgtgga ccgtgatggc aaggaccagg gtgtgaatgt
     1741 gcgggagaag gccaagcagc tggtggcttt gctgcgtgat gaggatcggc ttcgggagga
     1801 gcgcgcccac gcgctcaaga ccaaggagaa actggcacag actgccacgg cttcctcagc
     1861 agctgtgggc tctggaccac ctcccgaggc ggagcaggca tggccacaga gcagcgggga
     1921 ggaggagttg cagctccagc tggccctggc tatgagcaag gaggaggctg accagccccc
     1981 gtcctgcggc cccgaggatg acgtccagct ccagctggcc cttagtttga gccgagagga
     2041 gcacgataag gaagagcgga tccgtcgtgg ggatgacctg aggctgcaga tggcaataga
     2101 ggagagcaag agggagactg ggggcaagga ggagtcatct cttatggatc ttgctgacgt
     2161 cttcacaacc ccagcccctc cacaggcctc agacccttgg gggggcccgg catctgtgcc
     2221 tactgctgtc cctgtggctg ctgctgcctc agacccttgg ggggcacctg ctgtccctcc
     2281 agctgctgat ccttggggtg gtgcagcccc cacgccagcc tccggggatc cttggaggcc
     2341 tgcagcccct acagggccct ctgttgaccc ttggggtggg accccagctc ctgcagctgg
     2401 agaggggccc acatctgacc cgtggggaag cgctgatggt ggggcccctg tcagtggacc
     2461 cccatcttct gacccctggg cacctgcccc agctttctca gacccttggg gaggttcacc
     2521 tgcaaagccc agcagcaatg gcactgcagt tgggggcttt gacactgagc ccgatgagtt
     2581 ctcagacttc gaccgactcc gcactgccct gccaacctct gggagcagca caggggaatt
     2641 ggagttgctc gcaggggagg ttcctgctcg cagccccggg gcattcgaca tgagtggggt
     2701 tgggggatct ctggctgagt ctgtggggag tccccctcct gcggcaaccc caactcctac
     2761 cccgccaaca cggaagactc cggagtcatt cctaggccct aatgcagccc ttgtggactt
     2821 ggactcactg gtgagccgac caggccctac accgccagga gccaaggctt ccaacccatt
     2881 ccttccaagt ggagctccgg ccaccggccc ctccgtcacc aatcctttcc agccagcacc
     2941 ccctgcaacg ctcaccctga accagctccg tctcagtcct gtgcccccag ttcctggagc
     3001 gccacccacc tacatctctc ctcttggtgg aggccctggc ctgcccccca tgatgccccc
     3061 gggtccccca gcccccaaca ctactaaccc cttcctccta gtcgacggta ccgcgggccc
     3121 gggatccacc ggatctagat aactgatcat aatcagccat accacatttg tagaggtttt
     3181 acttgcttta aaaaacctcc cacacctccc cctgaacctg aaacataaaa tgaatgcaat
     3241 tgttgttgtt aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     3301 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     3361 caatgtatct taacgcgtaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt
     3421 tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat
     3481 caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat
     3541 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac
     3601 tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc
     3661 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga
     3721 gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca
     3781 cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcaggtg
     3841 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     3901 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     3961 agagtcctga ggcggaaaga accagctgtg gaatgtgtgt cagttagggt gtggaaagtc
     4021 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag
     4081 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     4141 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
     4201 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
     4261 ctcggcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg
     4321 caaagatcga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc
     4381 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
     4441 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
     4501 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca agacgaggca gcgcggctat
     4561 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
     4621 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg
     4681 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
     4741 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
     4801 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
     4861 ccgaactgtt cgccaggctc aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc
     4921 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg
     4981 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata
     5041 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg
     5101 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac
     5161 tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc
     5221 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     5281 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccctaggg ggaggctaac
     5341 tgaaacacgg aaggagacaa taccggaagg aacccgcgct atgacggcaa taaaaagaca
     5401 gaataaaacg cacggtgttg ggtcgtttgt tcataaacgc ggggttcggt cccagggctg
     5461 gcactctgtc gataccccac cgagacccca ttggggccaa tacgcccgcg tttcttcctt
     5521 ttccccaccc caccccccaa gttcgggtga aggcccaggg ctcgcagcca acgtcggggc
     5581 ggcaggccct gccatagcct caggttactc atatatactt tagattgatt taaaacttca
     5641 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     5701 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     5761 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     5821 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     5881 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     5941 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     6001 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     6061 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     6121 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     6181 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     6241 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     6301 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     6361 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     6421 gttatcccct gattctgtgg ataaccgtat taccgc
//
