LOCUS       sequence-477136-        4732 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses EGFP-lifectin fusion in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4732)
  AUTHORS   Zhang H, Lesnov GD, Subach OM, Zhang W, Kuzmicheva TP, Vlaskina AV, 
            Samygina VR, Chen L, Ye X, Nikolaeva AY, Gabdulkhakov A, Papadaki S,
            Qin W, Borshchevskiy V, Perfilov MM, Gavrikov AS, Drobizhev M, 
            Mishin AS, Piatkevich KD, Subach FV
  TITLE     Bright and stable monomeric green fluorescent protein derived from 
            StayGold.
  JOURNAL   Nat Methods. 2024 Apr;21(4):657-665. doi: 
            10.1038/s41592-024-02203-y. Epub 2024 Feb 26.
  PUBMED    38409224
REFERENCE   2  (bases 1 to 4732)
  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"; doi: 
            "10.1038/s41592-024-02203-y"; journalName: "Nat Methods"; date: 
            "2024-04"; volume: "21"; issue: "4"; pages: "657-665"
COMMENT     Sequence Label: EGFP-lifeactin
FEATURES             Location/Qualifiers
     source          1..4732
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             join(34..36,37..750)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(34..36,37..747)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(76..97)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(337..356)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     684..705
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    873..994
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(910..929)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     964..983
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1001..1456)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1138..1159)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1350..1369
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1483..1587
                     /gene="bla"
                     /label=AmpR promoter
     promoter        1589..1946
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1589..1780
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1612..1632)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      1797..1932
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             1981..2775
                     /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             1981..2775
                     /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(2035..2054)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2645..2664
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(2963..2982)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3007..3054
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      3383..3971
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3872..3891
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        4146..4449
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4450..4653
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4604..4624
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             4682..4732
                     /codon_start=1
                     /product="peptide that binds filamentous actin (Riedl et 
                     al., 2008)"
                     /label=Lifeact
                     /note="first 17 amino acids of budding yeast Abp140"
                     /translation="MGVADLIKKFESISKEE"
ORIGIN
        1 atccattcgt tgggggatcc accggtcgcc accgtgagca agggcgagga gctgttcacc
       61 ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg
      121 tccggcgagg gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc
      181 accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag
      241 tgcttcagcc gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc
      301 gaaggctacg tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc
      361 gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac
      421 ttcaaggagg acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac
      481 gtctatatca tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac
      541 aacatcgagg acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc
      601 gacggccccg tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa
      661 gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc
      721 actctcggca tggacgagct gtacaagtaa agcggccgcg actctagatc ataatcagcc
      781 ataccacatt tgtagaggtt ttacttgctt taaaaaacct cccacacctc cccctgaacc
      841 tgaaacataa aatgaatgca attgttgttg ttaacttgtt tattgcagct tataatggtt
      901 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
      961 gttgtggttt gtccaaactc atcaatgtat cttaaggcgt aaattgtaag cgttaatatt
     1021 ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa
     1081 atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca
     1141 gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc
     1201 gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg
     1261 aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg
     1321 ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg
     1381 gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg
     1441 ccgctacagg gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     1501 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     1561 cttcaataat attgaaaaag gaagagtcct gaggcggaaa gaaccagctg tggaatgtgt
     1621 gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     1681 atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta
     1741 tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc
     1801 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     1861 tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct
     1921 tttttggagg cctaggcttt tgcaaagatc gatcaagaga caggatgagg atcgtttcgc
     1981 atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc
     2041 ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca
     2101 gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg
     2161 caagacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg
     2221 ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag
     2281 gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg
     2341 cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc
     2401 atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa
     2461 gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgag catgcccgac
     2521 ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat
     2581 ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac
     2641 atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc
     2701 ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt
     2761 gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg acgcccaacc
     2821 tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg
     2881 ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg
     2941 cccaccctag ggggaggcta actgaaacac ggaaggagac aataccggaa ggaacccgcg
     3001 ctatgacggc aataaaaaga cagaataaaa cgcacggtgt tgggtcgttt gttcataaac
     3061 gcggggttcg gtcccagggc tggcactctg tcgatacccc accgagaccc cattggggcc
     3121 aatacgcccg cgtttcttcc ttttccccac cccacccccc aagttcgggt gaaggcccag
     3181 ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc ctcaggttac tcatatatac
     3241 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     3301 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     3361 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     3421 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     3481 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     3541 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     3601 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     3661 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     3721 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     3781 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     3841 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     3901 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     3961 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     4021 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcca
     4081 tgcattagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
     4141 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
     4201 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
     4261 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
     4321 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
     4381 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
     4441 attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca
     4501 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
     4561 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg
     4621 cgtgtacggt gggaggtcta tataagcaga gctggtttag tgaaccgtca gatccgctag
     4681 catgggcgtg gccgacctga tcaagaagtt cgagagcatc agcaaggagg ag
//
