LOCUS       sequence-296784-        6976 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Detection of Dengue virus infection in live cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6976)
  AUTHORS   Medin CL, Valois S, Patkar CG, Rothman AL
  TITLE     A plasmid-based reporter system for live cell imaging of dengue 
            virus infected cells.
  JOURNAL   J Virol Methods. 2015 Jan;211:55-62. doi: 
            10.1016/j.jviromet.2014.10.010. Epub 2014 Oct 28.
  PUBMED    25445884
REFERENCE   2  (bases 1 to 6976)
  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: "J Virol 
            Methods."; date: "2015-01"; volume: "211:55-62. doi"; pages: " 
            10.1016/j.jviromet.2014.10.010. Epub 2014 Oct 28"
COMMENT     Sequence Label: p4B5-EGFP
FEATURES             Location/Qualifiers
     source          1..6976
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      674..683
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1475..1495
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(1499..1501,1502..2215)
                     /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(1499..1501,1502..2212)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1541..1562)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1802..1821)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2149..2170
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2240..2281
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2291..2308
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2331..2348)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2337..2561
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2607..3035
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2694..2713)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2904..2925
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3044..3064)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3049..3378
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3229..3364
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3291..3310
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3445..4239
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3499..3518)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4109..4128
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4415..4536
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4452..4471)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4506..4525
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4585..4601)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4585..4601)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4598..4620)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4609..4625
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4633..4639,4640..4657,4658..4663))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4678..4699
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4816..4833)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4987..5575)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5067..5086)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5746..6537,6538..6606))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6369..6388
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6607..6711)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6786..6805)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctg
      661 gctagttaag cttgccacca tggcgaacga gatgggtttc ctagaaaaaa cgaagaaaga
      721 tctcggattg ggaagcatta caacccagca acccgagagc aacatcctgg acatagatct
      781 acgtcccgca tcagcatgga cgctgtatgc tgtggccaca acatttgtca caccaatgtt
      841 gagacacagc attgaaaatt cctcagtgaa cgtgtcccta acagctattg ccaaccaagc
      901 cacagtgtta atgggtcttg ggaaaggatg gccattgtca aagatggaca tcggagttcc
      961 ccttctcgcc attggatgct actcacaagt caaccccata actctcacag cagctctttt
     1021 cttactggta gcacattatg ccatcatagg gccaggactc caagcaaaag caaccagaga
     1081 agctcagaaa agagcagcag cgggcatcat gaaaaaccca actgtcgatg gaataacagt
     1141 gattgaccta gatccaatac cctatgatcc aaagtttgaa aagcagttgg gacaagtaat
     1201 gctcctagtc ctctgcgtga ctcaagtgtt gatgatgagg actacatggg ctctgtgtga
     1261 ggctttaacc ttagcgaccg ggcctatctc cacattgtgg gaaggaaatc cagggaggtt
     1321 ttggaacact accattgcag tgtcaatggc taacattttt agagggagtt acttggccgg
     1381 agctggactt ctcttttcca tcatgaagaa cacaaccaac acgagaaggg gaactggcaa
     1441 cataggagag acgcttggag aattcgccac catgccgaag aaaaagcgga aggttggcgt
     1501 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
     1561 cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa
     1621 gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt
     1681 gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca
     1741 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa
     1801 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
     1861 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
     1921 ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat
     1981 caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca
     2041 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     2101 gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     2161 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca agtaactcga
     2221 gtctagaggg cccttcgaag gtaagcctat ccctaaccct ctcctcggtc tcgattctac
     2281 gcgtaccggt catcatcacc atcaccattg agtttaaacc cgctgatcag cctcgactgt
     2341 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     2401 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag
     2461 taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga
     2521 agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac
     2581 cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg
     2641 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     2701 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     2761 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     2821 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     2881 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     2941 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     3001 aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtggaatgt gtgtcagtta
     3061 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     3121 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     3181 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     3241 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     3301 gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     3361 ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg atctgatcaa
     3421 gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg
     3481 gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat cggctgctct
     3541 gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac
     3601 ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg
     3661 acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg
     3721 ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa
     3781 gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca
     3841 ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt
     3901 gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc
     3961 aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc
     4021 ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg
     4081 ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt
     4141 ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag
     4201 cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgcga
     4261 aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt
     4321 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg
     4381 cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg
     4441 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     4501 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc
     4561 tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
     4621 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
     4681 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     4741 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     4801 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     4861 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     4921 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     4981 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     5041 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     5101 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     5161 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     5221 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     5281 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     5341 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     5401 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     5461 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     5521 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     5581 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     5641 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     5701 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     5761 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     5821 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     5881 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     5941 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     6001 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     6061 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     6121 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     6181 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     6241 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     6301 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     6361 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     6421 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     6481 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     6541 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     6601 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     6661 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     6721 ccgaaaagtg ccacctgacg tcgacggatc gggagatctc ccgatcccct atggtgcact
     6781 ctcagtacaa tctgctctga tgccgcatag ttaagccagt atctgctccc tgcttgtgtg
     6841 ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta caacaaggca aggcttgacc
     6901 gacaattgca tgaagaatct gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg
     6961 gccagatata cgcgtt
//
