LOCUS       sequence-335127-        7454 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of EPS15-GFP-His..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7454)
  AUTHORS   Prasai B, Haber GJ, Strub MP, Ahn R, Ciemniecki JA, Sochacki KA, 
            Taraska JW
  TITLE     The nanoscale molecular morphology of docked exocytic dense-core 
            vesicles in neuroendocrine cells.
  JOURNAL   Nat Commun. 2021 Jun 25;12(1):3970. doi: 10.1038/s41467-021-24167-9.
  PUBMED    34172739
REFERENCE   2  (bases 1 to 7454)
  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 
            Commun."; date: "2021-06-25"; volume: "12(1):3970. doi"; pages: " 
            10.1038/s41467-021-24167-9"
COMMENT     Sequence Label: EPS15-GFP-His
FEATURES             Location/Qualifiers
     source          1..7454
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      3367..3376
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3373..3375,3376..3378,3379..4089)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3418..3439)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3679..3698)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4026..4047
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             4099..4116
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    4242..4363
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4279..4298)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4333..4352
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4370..4825)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4507..4528)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4719..4738
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4852..4956
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4958..5315
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4958..5149
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4981..5001)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5166..5301
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5350..6144
                     /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             5350..6144
                     /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(5404..5423)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6014..6033
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6332..6351)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6376..6423
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6752..7340
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7241..7260
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgagac catggctgcg gcggcccagc tctccctgac acagttatca
      661 agtgggaatc ctgtatatga aaaatactat agacaggttg atacaggcaa tactggaagg
      721 gtgttggctt ctgatgctgc tgctttcctg aaaaaatcag ggcttccaga cttgatactt
      781 ggaaagattt gggatttagc cgacacagat ggcaaaggta tcctgaacaa acaagaattc
      841 tttgttgctt tgcgtcttgt ggcatgtgcc cagaatggat tggaagtttc actaagtagt
      901 ttgaacctgg ctgttcctcc accaagattt catgatacca gtagtccttt gctaatcagt
      961 ggaacctctg cagctgagct cccatgggct gtaaaacctg aagataaggc caaatatgat
     1021 gcaatatttg atagtttaag cccagtgaat ggatttctgt ctggtgataa agtgaaacca
     1081 gtgttgctca actctaagtt acctgtggat atccttggaa gagtttggga gttgagtgat
     1141 attgaccatg atggaatgct tgacagagat gagtttgcag ttgccatgtt tttggtatac
     1201 tgtgcactgg agaaagaacc tgtgccaatg tccttgcctc cagccttggt gccaccatct
     1261 aagagaaaaa cgtgggttgt atcccctgca gaaaaagcta aatatgatga aatcttcctg
     1321 aaaactgata aagatatgga cggatttgtg tctggattgg aggtccgtga aatattcttg
     1381 aaaacaggtt taccttctac cttactagcc catatatggt cattatgcga cacaaaggac
     1441 tgtgggaagc tttcaaagga tcagtttgcc ttggcttttc acttaatcag tcagaagtta
     1501 atcaagggca ttgatcctcc tcacgttctt actcctgaaa tgattccacc atcagacagg
     1561 gccagtttac aaaagaacat cataggatca agtcctgttg cagatttctc tgctattaag
     1621 gaactagata ctcttaacaa tgaaatagtt gacctacaga gggaaaagaa taatgtggaa
     1681 caggacctta aggagaagga agatactatt aaacagagga caagtgaggt tcaggatctt
     1741 caagatgaag ttcaaaggga gaatactaat ctgcaaaaac tacaggccca gaaacagcag
     1801 gtacaggaac tccttgatga actggatgag cagaaagccc agctggagga gcaactcaag
     1861 gaagtcagaa agaaatgtgc tgaggaggcc caactgatct cttctctgaa agctgaatta
     1921 actagtcagg aatcgcagat ctccacttac gaagaagaat tggcaaaagc tagagaagag
     1981 ctgagccgtc tacagcaaga aacagcagaa ttggaggaga gtgtagagtc agggaaggct
     2041 cagttggaac ctcttcagca gcacctacaa gattcacaac aggaaattag ttcaatgcaa
     2101 atgaaactga tggaaatgaa agatttggaa aatcataata gtcagttaaa ttggtgcagt
     2161 agcccacaca gcattcttgt aaacggagct acagattatt gcagcctcag caccagcagc
     2221 agtgaaacag ccaaccttaa tgaacatgtt gaaggccaga gcaacctaga gtctgagccc
     2281 atacaccagg aatctccagc aagaagtagt cctgaactac tgccttctgg tgtgactgat
     2341 gaaaatgagg tgactacagc tgttactgaa aaagtttgtt ctgaactcga caataataga
     2401 cattcaaaag aggaagatcc atttaatgta gactcaagtt cgctgacagg tccagttgca
     2461 gatacaaact tggatttttt ccagtctgat ccttttgttg gcagtgatcc tttcaaggat
     2521 gatccttttg gaaaaatcga tccatttggt ggtgatcctt tcaaaggttc agatccattt
     2581 gcatcagact gtttcttcag gcaatctact gatccttttg ccacttcaag cactgaccct
     2641 ttcagtgcag ccaacaatag cagtattaca tcggtagaaa cgttgaagca caatgatcct
     2701 tttgctcctg gtggaacagt tgttgcagca agcgattcag ccacagaccc ctttgcttct
     2761 gtttttggga atgaatcatt tggaggtgga tttgctgact tcagcacatt gtcaaaggtc
     2821 aacaatgaag atccttttcg ttcagccaca tcgagctctg tcagcaacgt agtgattaca
     2881 aaaaatgtat ttgaggaaac atcggtcaaa agtgaagatg aacccccagc actgccacca
     2941 aagatcggaa ctccaacaag accctgccct ctaccacctg ggaaaagatc catcaacaaa
     3001 ttggattctc ctgatccctt taaactgaat gatccatttc agcctttccc aggcaacgat
     3061 agccccaaag aaaaagatcc tgaaatattt tgtgatccat tcacttctgc tactaccact
     3121 accaataaag aggctgatcc aagcaatttt gccaacttca gtgcttatcc ctctgaagaa
     3181 gatatgatcg aatgggccaa gagggaaagt gagagagagg aagagcagag gcttgcccga
     3241 ctaaatcagc aggaacaaga agacttagaa ctggctattg cactcagcaa atctgagata
     3301 tcagaagcac ggatgggttc gacctcgggt tcgaccaagg gtcagctggc ccgggatcca
     3361 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
     3421 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
     3481 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
     3541 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
     3601 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
     3661 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
     3721 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     3781 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     3841 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     3901 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     3961 cccgacaacc actacctgag cacccagtcc aagctgagca aagaccccaa cgagaagcgc
     4021 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     4081 ctgtacaagg gcagcagcca tcatcatcat catcactaaa gcggccgcga ctctagatca
     4141 taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc ccacacctcc
     4201 ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt attgcagctt
     4261 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     4321 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttaaggcgta aattgtaagc
     4381 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     4441 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
     4501 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
     4561 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt
     4621 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
     4681 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
     4741 ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
     4801 cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc ggggaaatgt gcgcggaacc
     4861 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     4921 tgataaatgc ttcaataata ttgaaaaagg aagagtcctg aggcggaaag aaccagctgt
     4981 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     5041 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     5101 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     5161 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     5221 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
     5281 gaggaggctt ttttggaggc ctaggctttt gcaaagatcg atcaagagac aggatgagga
     5341 tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
     5401 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
     5461 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
     5521 aatgaactgc aagacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
     5581 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
     5641 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
     5701 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     5761 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     5821 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgagc
     5881 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     5941 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
     6001 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
     6061 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
     6121 cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga
     6181 cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct
     6241 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg
     6301 agttcttcgc ccaccctagg gggaggctaa ctgaaacacg gaaggagaca ataccggaag
     6361 gaacccgcgc tatgacggca ataaaaagac agaataaaac gcacggtgtt gggtcgtttg
     6421 ttcataaacg cggggttcgg tcccagggct ggcactctgt cgatacccca ccgagacccc
     6481 attggggcca atacgcccgc gtttcttcct tttccccacc ccacccccca agttcgggtg
     6541 aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc tgccatagcc tcaggttact
     6601 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     6661 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     6721 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     6781 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     6841 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     6901 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     6961 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     7021 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     7081 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     7141 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     7201 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     7261 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     7321 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     7381 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     7441 ttaccgccat gcat
//
