LOCUS       sequence-47288-e        4825 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 4825)
  AUTHORS   Suzuki T, Arai S, Takeuchi M, Sakurai C, Ebana H, Higashi T, 
            Hashimoto H, Hatsuzawa K, Wada I
  TITLE     Development of cysteine-free fluorescent proteins for the oxidative 
            environment.
  JOURNAL   PLoS One. 2012;7(5):e37551. Epub 2012 May 23.
  PUBMED    22649538
REFERENCE   2  (bases 1 to 4825)
  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: "PLoS One.";
            date: "2012"; volume: "7(5)"; pages: "e37551. Epub 2012 May 23"
COMMENT     Sequence Label: ss-cfSGFP2
FEATURES             Location/Qualifiers
     source          1..4825
                     /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      765..774
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(771..773,774..776,777..1487)
                     /codon_start=1
                     /product="strongly enhanced green fluorescent protein 
                     (Kremers et al., 2007)"
                     /label=SGFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFISTTGKLPVPWPTLVTTLTYGVQMFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(816..837)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1077..1096)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1424..1445
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1613..1734
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1650..1669)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1704..1723
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1741..2196)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1878..1899)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2090..2109
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2223..2327
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2329..2686
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2329..2520
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2352..2372)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2537..2672
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2721..3515
                     /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             2721..3515
                     /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(2775..2794)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3385..3404
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3703..3722)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3747..3794
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4123..4711
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4612..4631
                     /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 atctacaatg ccgtcttctg tctcgtgggg catcctcctg ctggcaggcc
      661 tgtgctgcct ggtccctgtc tccctggctg aggatcccca gggagatgct gcccagaaga
      721 gaattctgca gtcgacggta ccgcgggccc gggatccacc ggtcgccacc atggtgagca
      781 agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa
      841 acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac ggcaagctga
      901 ccctgaagtt catctccacc accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca
      961 ccctgaccta cggcgtgcag atgttcgccc gctaccccga ccacatgaag cagcacgact
     1021 tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc ttcaaggacg
     1081 acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg gtgaaccgca
     1141 tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac aagctggagt
     1201 acaactacaa cagccacaac gtctatatca cggccgacaa gcagaagaac ggcatcaagg
     1261 ccaacttcaa gatccgccac aacatcgagg acggcggcgt gcagctcgcc gaccactacc
     1321 agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac tacctgagca
     1381 cccagtccaa actgagcaaa gaccccaacg agaagcgcga tcacatggtc ctgctggagt
     1441 tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagtaa agcggccgcg
     1501 actctagatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt taaaaaacct
     1561 cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt
     1621 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     1681 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttaaggcgt
     1741 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
     1801 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
     1861 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
     1921 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
     1981 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
     2041 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
     2101 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
     2161 cacccgccgc gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt cggggaaatg
     2221 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     2281 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtcct gaggcggaaa
     2341 gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg
     2401 cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg
     2461 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc
     2521 gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca
     2581 tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt
     2641 ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaagatc gatcaagaga
     2701 caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg
     2761 cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg
     2821 ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt
     2881 ccggtgccct gaatgaactg caagacgagg cagcgcggct atcgtggctg gccacgacgg
     2941 gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat
     3001 tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat
     3061 ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg
     3121 accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg
     3181 atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc
     3241 tcaaggcgag catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc
     3301 cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg
     3361 tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg
     3421 gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca
     3481 tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac
     3541 cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga
     3601 aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga
     3661 tctcatgctg gagttcttcg cccaccctag ggggaggcta actgaaacac ggaaggagac
     3721 aataccggaa ggaacccgcg ctatgacggc aataaaaaga cagaataaaa cgcacggtgt
     3781 tgggtcgttt gttcataaac gcggggttcg gtcccagggc tggcactctg tcgatacccc
     3841 accgagaccc cattggggcc aatacgcccg cgtttcttcc ttttccccac cccacccccc
     3901 aagttcgggt gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc
     3961 ctcaggttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     4021 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     4081 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     4141 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     4201 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     4261 aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     4321 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     4381 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     4441 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     4501 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     4561 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     4621 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     4681 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     4741 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     4801 ggataaccgt attaccgcca tgcat
//
