LOCUS       sequence-324365-        4962 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression and localization.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4962)
  AUTHORS   Miserey-Lenkei S, Trajkovic K, D'Ambrosio JM, Patel AJ, Copic A, 
            Mathur P, Schauer K, Goud B, Albanese V, Gautier R, Subra M, Kovacs 
            D, Barelli H, Antonny B
  TITLE     A comprehensive library of fluorescent constructs of SARS-CoV-2 
            proteins and their initial characterisation in different cell types.
  JOURNAL   Biol Cell. 2021 Jul;113(7):311-328. doi: 10.1111/boc.202000158. Epub
            2021 May 10.
  PUBMED    33666950
REFERENCE   2  (bases 1 to 4962)
  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: 
            "10.1111/boc.202000158"; journalName: "Biol Cell"; date: "2021-07"; 
            volume: "113"; issue: "7"; pages: "311-328"
COMMENT     Sequence Label: EGFP-Nsp7
FEATURES             Location/Qualifiers
     source          1..4962
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..204
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     155..175
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      243..252
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(249..251,252..254,255..965)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(249..251,252..254,255..965)
                     /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(294..315)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(555..574)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     902..923
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1386..1507
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1423..1442)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1477..1496
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1514..1969)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1651..1672)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1863..1882
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1996..2100
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2102..2459
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2102..2293
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2125..2145)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2310..2445
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2494..3288
                     /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             2494..3288
                     /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(2548..2567)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3158..3177
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3476..3495)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3520..3567
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      3896..4484
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4385..4404
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        4659..4962
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
       61 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
      121 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
      181 tgggaggtct atataagcag agctggttta gtgaaccgtc agatccgcta gcgctaccgg
      241 tcgccaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
      301 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg
      361 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
      421 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc
      481 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
      541 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
      601 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
      661 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc
      721 agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc
      781 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
      841 acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc
      901 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt
      961 acaagtccgg actcagatct cgagctcaag cttcgaattc tatgagcaag atgagcgacg
     1021 taaaatgcac gtcagttgtt ctcttgagcg tcctgcaaca actcagagtg gagtctagtt
     1081 ccaagctctg ggctcaatgc gttcagcttc acaatgacat tctgttggcg aaagatacca
     1141 cggaggcatt cgagaagatg gtctcacttc tgagcgttct cctgtcaatg caaggggcgg
     1201 tcgatatcaa taaattgtgt gaggaaatgc tcgacaaccg ggcgacattg cagtaaggat
     1261 ccaccggatc tagataactg atcataatca gccataccac atttgtagag gttttacttg
     1321 ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg
     1381 ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     1441 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     1501 tatcttaacg cgtaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt
     1561 taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa
     1621 gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag
     1681 aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt
     1741 gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac
     1801 cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag
     1861 gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg
     1921 cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc aggtggcact
     1981 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     2041 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     2101 cctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
     2161 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
     2221 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     2281 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
     2341 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg
     2401 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaag
     2461 atcgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca
     2521 ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc
     2581 ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc
     2641 aagaccgacc tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg
     2701 ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg
     2761 gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct
     2821 gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct
     2881 acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa
     2941 gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa
     3001 ctgttcgcca ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt gacccatggc
     3061 gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt
     3121 ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct
     3181 gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc
     3241 gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg
     3301 ggttcgaaat gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg
     3361 ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc
     3421 tccagcgcgg ggatctcatg ctggagttct tcgcccaccc tagggggagg ctaactgaaa
     3481 cacggaagga gacaataccg gaaggaaccc gcgctatgac ggcaataaaa agacagaata
     3541 aaacgcacgg tgttgggtcg tttgttcata aacgcggggt tcggtcccag ggctggcact
     3601 ctgtcgatac cccaccgaga ccccattggg gccaatacgc ccgcgtttct tccttttccc
     3661 caccccaccc cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag
     3721 gccctgccat agcctcaggt tactcatata tactttagat tgatttaaaa cttcattttt
     3781 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
     3841 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     3901 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     3961 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     4021 gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga
     4081 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     4141 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     4201 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     4261 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     4321 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     4381 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     4441 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     4501 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat
     4561 cccctgattc tgtggataac cgtattaccg ccatgcatta gttattaata gtaatcaatt
     4621 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
     4681 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
     4741 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
     4801 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
     4861 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
     4921 acttggcagt acatctacgt attagtcatc gctattacca tg
//
