LOCUS       sequence-170487-        7906 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 7906)
  AUTHORS   Yamada KM
  TITLE     Yamada Lab Plasmids
  JOURNAL   unpublished
REFERENCE   2  (bases 1 to 7906)
  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: 
            "unpublished"
COMMENT     Sequence Label: pEGFP Vinculin
FEATURES             Location/Qualifiers
     source          1..7906
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     130..150
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      218..227
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(224..226,227..229,230..940)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(224..226,227..229,230..940)
                     /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(269..290)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(530..549)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     877..898
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    4305..4426
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4342..4361)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4396..4415
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4433..4888)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4570..4591)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4782..4801
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4915..5019
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5021..5378
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5021..5212
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5044..5064)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5229..5364
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5413..6207
                     /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             5413..6207
                     /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(5467..5486)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6077..6096
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6395..6414)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6439..6486
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6815..7403
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7304..7323
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        7578..7881
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        join(7882..7906,1..179)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
ORIGIN
        1 atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca
       61 atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg
      121 ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagagctg
      181 gtttagtgaa ccgtcagatc cgctagcgct accggtcgcc accatggtga gcaagggcga
      241 ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca
      301 caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa
      361 gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac
      421 ctacggcgtg cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa
      481 gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa
      541 ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct
      601 gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta
      661 caacagccac aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt
      721 caagatccgc cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa
      781 cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc
      841 cgccctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac
      901 cgccgccggg atcactctcg gcatggacga gctgtacaag tccggactca gatctcgagc
      961 tgcggccgcc ggatccaagc ttatgcccgt cttccacacg cgcaccatcg agagcatctt
     1021 ggagcccgtg gctcagcaga tctcccacct ggtcatcatg cacgaggagg gggaggtaga
     1081 cggcaaggcc atcccggacc tcaccgcccc cgtgtcggcc gtgcaggccg ctgtcagcaa
     1141 cctggtgcgg gttggaaaag aaactgtgca gacaacagaa gaccagatct tgaaaaggga
     1201 tatgccacca gcattcatca aagtagagaa tgcctgcacc aagctcgttc gagcagccca
     1261 gatgctgcaa gcagatcctt attcagtacc agctcgtgac tacctaattg atggatcaag
     1321 aggcatcctt tctggaacat cagacttact tctgacattt gatgaagcgg aggtccgtaa
     1381 aatcatccgt gtctgcaaag gaatattgga atatctgact gtggcagaag tagtagagac
     1441 tatggaggat ttggtgacat atacaaagaa tctagggcca ggaatgacaa agatggcgaa
     1501 aatgatcgat gagagacaac aggaattaac tcatcaggaa catagggtta tgctggtgaa
     1561 ctccatgaat actgtgaagg agctattgcc tgtacttatt tcagctatga agatctttgt
     1621 aaccacaaaa aacactaaaa gccagggaat agaagaggcc ttgaaaaatc gcaatttcac
     1681 agtagagaaa atgagtgctg agataaatga aataatccgt gtattacaac tcacttcctg
     1741 ggatgaagat gcctgggcca gcaaggacac tgaagccatg aaaagagctt tagccctaat
     1801 agattcaaag atgaaccagg caaaaggctg gctgagagat ccaaacgcac ctccagggga
     1861 tgctggtgag caagcaataa ggcagatcct tgatgaagct ggaaaagcag gagaattgtg
     1921 tgcaggcaaa gaacgcaggg agattctggg aacatgcaaa actttgggcc aaatgactga
     1981 tcaacttgct gatctccgag ctagaggaca gggtgctaca ccgatggcaa tgcagaaggc
     2041 acagcaggtg tcacaaggcc tggatttgct cactgcaaaa gtggagaatg cagcccggaa
     2101 attggaggcc atgacaaact ctaagcaggc tattgcaaag aagattgatg ctgctcagaa
     2161 ctggcttgcg gatcccaacg gcggaagtga aggagaagaa cacattcgag gaattatgtc
     2221 tgaagcaagg aaagttgcag aattgtgtga ggagcctaaa gaaagagatg atatccttcg
     2281 ctccttgggg gaaatctctg ctctgacagc taagctgtca gatctgcgac gacatgggaa
     2341 aggcgactct cctgaggccc gtgcattggc caagcaaata gctacatcac ttcagaactt
     2401 acagtccaaa acaaacagag ctgtagcaaa tactaggcca gttaaagctg ctgtccattt
     2461 ggagggcaag attgagcaag cccagaggtg gatagataat cctacagttg atgatcgggg
     2521 agtaggccag gcagcaattc ggggtttggt tgcagaaggt cgtcgtctag ccaatgtcat
     2581 gatgggacct tatcgtcagg acctgcttgc caaatgtgac cgtgtagacc agctggctgc
     2641 tcagcttgct gaccttgcag caagagggga aggagagtct cctcaggcta gggcaattgc
     2701 tgctcagctt caggactccc tgaaggatct caaagcacgg atgcaagaag caatgaccca
     2761 ggaggtgtct gatgttttca gtgacactac aacccctatt aagttgttag cagtagcagc
     2821 cactgctcct tctgatactc ccaatagaga agaggtgttt gaagaaagag cagcaaattt
     2881 tgaaaaccat gctgctagac tgggagcaac agcagaaaaa gcagctgcag ttggaactgc
     2941 taataaaact actgtggaag gcattcaggc aacagtcaaa tctgcaaggg agcttacacc
     3001 acaggtagta tcggctgctc gaatcctcct gagaaatcct ggaaatcaag ctgcttatga
     3061 gcattttgag acaatgaaaa accaatggat tgataatgta gaaaagatga cagggctggt
     3121 ggatgaggcc atcgatacca agtctctgtt ggatgcatca gaagaggcta ttaagaagga
     3181 tcttgataaa tgtaaagttg caatggccaa tatgcaacct cagatgctgg tagctggagc
     3241 caccagcatt gctagacgag cgaaccgcat cctgcttgtg gcaaaacggg aggttgaaaa
     3301 ttcagaagac cctaaattca gggaggctgt taaagcagct tctgatgagc tgagcaaaac
     3361 catatcaccg atggtaatgg atgctaaagc tgtagcagga aatatctctg atcctggttt
     3421 gcagaagagt ttcttggatt ctggatacag gattctggga gctgtggcca aagtcagaga
     3481 agcctttcag cctcaggagc cagactttcc ccctcctcct cctgacctag agcatctcca
     3541 tctgactgat gagcttgctc ctccaaaacc accacttcca gaaggtgagg ttcccccacc
     3601 cagaccacca ccccctgaag aaaaagatga ggagttccca gagcagaaag caggagaagc
     3661 tattaatcag cccatgatga tggctgctag gcagttgcat gacgaggctc ggaaatggtc
     3721 tagcaagggt aacgacatca ttgctgctgc taaacgaatg gcgctgctca tggctgagat
     3781 gtcacgcctg gtgcgaggag gcagcggaaa caagcgtgcc ctcatccagt gtgcaaaaga
     3841 tattgctaag gcatcggatg aagtcactcg attggccaaa gaggtggcca agcaatgtac
     3901 tgataaacgc attagaacaa acctcttaca ggtctgtgag cgaatcccaa ccatcagcac
     3961 gcagctcaaa attctttcca cagtgaaagc taccatgctg ggcaggacta acatcagcga
     4021 tgaagaatca gaacaggcaa ctgagatgtt ggttcataac gcccagaacc tcatgcagtc
     4081 tgtgaaggaa actgtgagag aagctgaagc agcatccatt aagataagaa cagatgccgg
     4141 attcactctg cgctgggtca gaaagacccc atggtatcag taaacactct agataactga
     4201 tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac ctcccacacc
     4261 tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag
     4321 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     4381 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttaaggc gtaaattgta
     4441 agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac
     4501 caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg
     4561 agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa
     4621 gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt
     4681 tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt
     4741 agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga
     4801 gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc
     4861 gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga
     4921 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     4981 ccctgataaa tgcttcaata atattgaaaa aggaagagtc ctgaggcgga aagaaccagc
     5041 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     5101 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     5161 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     5221 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     5281 taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt
     5341 agtgaggagg cttttttgga ggcctaggct tttgcaaaga tcgatcaaga gacaggatga
     5401 ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg
     5461 gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg
     5521 ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc
     5581 ctgaatgaac tgcaagacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct
     5641 tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa
     5701 gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg
     5761 gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa
     5821 gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat
     5881 gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg
     5941 agcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc
     6001 atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac
     6061 cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg
     6121 gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc
     6181 tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag
     6241 cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg
     6301 gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc
     6361 tggagttctt cgcccaccct agggggaggc taactgaaac acggaaggag acaataccgg
     6421 aaggaacccg cgctatgacg gcaataaaaa gacagaataa aacgcacggt gttgggtcgt
     6481 ttgttcataa acgcggggtt cggtcccagg gctggcactc tgtcgatacc ccaccgagac
     6541 cccattgggg ccaatacgcc cgcgtttctt ccttttcccc accccacccc ccaagttcgg
     6601 gtgaaggccc agggctcgca gccaacgtcg gggcggcagg ccctgccata gcctcaggtt
     6661 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     6721 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     6781 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     6841 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     6901 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     6961 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     7021 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     7081 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     7141 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     7201 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     7261 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     7321 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     7381 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     7441 tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc
     7501 gtattaccgc catgcattag ttattaatag taatcaatta cggggtcatt agttcatagc
     7561 ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc
     7621 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg
     7681 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat
     7741 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc
     7801 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta
     7861 ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatca
//
