LOCUS       sequence-321576-        6366 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses both eGFP and firefly luciferase in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6366)
  TITLE     Expression plasmid for production of murine IFN gamma
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6366)
  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            "Unpublished"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: eGFP-luciferase-C1
FEATURES             Location/Qualifiers
     source          1..6366
                     /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      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(613..615,616..618,619..1329)
                     /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(658..679)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(919..938)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1266..1287
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1372..3024
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     primer_bind     complement(1438..1456)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     polyA_signal    3154..3275
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3191..3210)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3245..3264
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3282..3737)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3419..3440)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3631..3650
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3764..3868
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3870..4227
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3870..4061
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3893..3913)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4078..4213
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4262..5056
                     /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             4262..5056
                     /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(4316..4335)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4926..4945
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5244..5263)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5288..5335
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5664..6252
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6153..6172
                     /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 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      661 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
      721 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
      781 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
      841 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      901 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
      961 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1021 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     1081 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     1141 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1201 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
     1261 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1321 ctgtacaagt ccggactcag atctcgagct caagcttcga attctgcagt tatggaagac
     1381 gccaaaaaca taaagaaagg cccggcgcca ttctatccgc tggaagatgg aaccgctgga
     1441 gagcaactgc ataaggctat gaagagatac gccctggttc ctggaacaat tgcttttaca
     1501 gatgcacata tcgaggtgga catcacttac gctgagtact tcgaaatgtc cgttcggttg
     1561 gcagaagcta tgaaacgata tgggctgaat acaaatcaca gaatcgtcgt atgcagtgaa
     1621 aactctcttc aattctttat gccggtgttg ggcgcgttat ttatcggagt tgcagttgcg
     1681 cccgcgaacg acatttataa tgaacgtgaa ttgctcaaca gtatgggcat ttcgcagcct
     1741 accgtggtgt tcgtttccaa aaaggggttg caaaaaattt tgaacgtgca aaaaaagctc
     1801 ccaatcatcc aaaaaattat tatcatggat tctaaaacgg attaccaggg atttcagtcg
     1861 atgtacacgt tcgtcacatc tcatctacct cccggtttta atgaatacga ttttgtgcca
     1921 gagtccttcg atagggacaa gacaattgca ctgatcatga actcctctgg atctactggt
     1981 ctgcctaaag gtgtcgctct gcctcataga actgcctgcg tgagattctc gcatgccaga
     2041 gatcctattt ttggcaatca aatcattccg gatactgcga ttttaagtgt tgttccattc
     2101 catcacggtt ttggaatgtt tactacactc ggatatttga tatgtggatt tcgagtcgtc
     2161 ttaatgtata gatttgaaga agagctgttt ctgaggagcc ttcaggatta caagattcaa
     2221 agtgcgctgc tggtgccaac cctattctcc ttcttcgcca aaagcactct gattgacaaa
     2281 tacgatttat ctaatttaca cgaaattgct tctggtggcg ctcccctctc taaggaagtc
     2341 ggggaagcgg ttgccaagag gttccatctg ccaggtatca ggcaaggata tgggctcact
     2401 gagactacat cagctattct gattacaccc gagggggatg ataaaccggg cgcggtcggt
     2461 aaagttgttc cattttttga agcgaaggtt gtggatctgg ataccgggaa aacgctgggc
     2521 gttaatcaaa gaggcgaact gtgtgtgaga ggtcctatga ttatgtccgg ttatgtaaac
     2581 aatccggaag cgaccaacgc cttgattgac aaggatggat ggctacattc tggagacata
     2641 gcttactggg acgaagacga acacttcttc atcgttgacc gcctgaagtc tctgattaag
     2701 tacaaaggct atcaggtggc tcccgctgaa ttggaatcca tcttgctcca acaccccaac
     2761 atcttcgacg caggtgtcgc aggtcttccc gacgatgacg ccggtgaact tcccgccgcc
     2821 gttgttgttt tggagcacgg aaagacgatg acggaaaaag agatcgtgga ttacgtcgcc
     2881 agtcaagtaa caaccgcgaa aaagttgcgc ggaggagttg tgtttgtgga cgaagtaccg
     2941 aaaggtctta ccggaaaact cgacgcaaga aaaatcagag agatcctcat aaaggccaag
     3001 aagggcggaa agatcgccgt gtaaggatcc accggatcta gataactgat cataatcagc
     3061 cataccacat ttgtagaggt tttacttgct ttaaaaaacc tcccacacct ccccctgaac
     3121 ctgaaacata aaatgaatgc aattgttgtt gttaacttgt ttattgcagc ttataatggt
     3181 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     3241 agttgtggtt tgtccaaact catcaatgta tcttaacgcg taaattgtaa gcgttaatat
     3301 tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc aataggccga
     3361 aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga gtgttgttcc
     3421 agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac
     3481 cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt ttttggggtc
     3541 gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta gagcttgacg
     3601 gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag
     3661 ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc
     3721 gccgctacag ggcgcgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     3781 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     3841 gcttcaataa tattgaaaaa ggaagagtcc tgaggcggaa agaaccagct gtggaatgtg
     3901 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     3961 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
     4021 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
     4081 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     4141 atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc
     4201 ttttttggag gcctaggctt ttgcaaagat cgatcaagag acaggatgag gatcgtttcg
     4261 catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt
     4321 cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc
     4381 agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact
     4441 gcaagacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt
     4501 gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca
     4561 ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat
     4621 gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg
     4681 catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga
     4741 agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcga gcatgcccga
     4801 cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa
     4861 tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga
     4921 catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt
     4981 cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct
     5041 tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc gacgcccaac
     5101 ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc
     5161 gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc
     5221 gcccacccta gggggaggct aactgaaaca cggaaggaga caataccgga aggaacccgc
     5281 gctatgacgg caataaaaag acagaataaa acgcacggtg ttgggtcgtt tgttcataaa
     5341 cgcggggttc ggtcccaggg ctggcactct gtcgataccc caccgagacc ccattggggc
     5401 caatacgccc gcgtttcttc cttttcccca ccccaccccc caagttcggg tgaaggccca
     5461 gggctcgcag ccaacgtcgg ggcggcaggc cctgccatag cctcaggtta ctcatatata
     5521 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     5581 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     5641 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     5701 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     5761 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
     5821 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     5881 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     5941 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     6001 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     6061 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     6121 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     6181 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     6241 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     6301 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     6361 atgcat
//
