LOCUS       sequence-271494-        6181 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the OgNLuc LumiFluor targeted to the mitochondira in 
            mammalian cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6181)
  AUTHORS   Parag-Sharma K, O'Banion CP, Henry EC, Musicant AM, Cleveland JL, 
            Lawrence DS, Amelio AL
  TITLE     Engineered BRET-Based Biologic Light Sources Enable Spatio-Temporal 
            Control Over Diverse Optogenetic Systems.
  JOURNAL   ACS Synth Biol. 2019 Dec 13. doi: 10.1021/acssynbio.9b00277.
  PUBMED    31834783
REFERENCE   2  (bases 1 to 6181)
  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: "ACS Synth 
            Biol. 2019 Dec 13. doi: 10.1021/acssynbio.9b00277."
COMMENT     Sequence Label: pEF1a-T2-OgNLuc
FEATURES             Location/Qualifiers
     source          1..6181
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          231..1173
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1130..1150
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      1228..1237
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1447..2151
                     /codon_start=1
                     /product="monomeric orange fluorescent protein with a large
                     Stokes shift (Shcherbakova et al., 2012)"
                     /label=LSSmOrange
                     /note="mammalian codon-optimized"
                     /translation="VSKGEENNMAIIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGF
                     QTVKLKVTKGGPLPFAWDILSPQFTYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGMEASSERMYPEDGALKG
                     EDKLRLKLKDGGHYTSEVKTTYKAKKPVQLPGAYIVDIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     1857..1876
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             2167..2682
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     polyA_signal    2805..2926
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2842..2861)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2896..2915
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2933..3388)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3070..3091)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3282..3301
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3415..3519
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3521..3878
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3521..3712
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3544..3564)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3729..3864
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3913..4707
                     /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             3913..4707
                     /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(3967..3986)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4577..4596
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4895..4914)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4939..4986
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5315..5903
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5804..5823
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        join(6181,1..1182)
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
ORIGIN
        1 gctccggtgc ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg
       61 gaggggtcgg caattgaacc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg
      121 atgtcgtgta ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag
      181 tagtcgccgt gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gtaagtgccg
      241 tgtgtggttc ccgcgggcct ggcctcttta cgggttatgg cccttgcgtg ccttgaatta
      301 cttccacgcc cctggctgca gtacgtgatt cttgatcccg agcttcgggt tggaagtggg
      361 tgggagagtt cgaggccttg cgcttaagga gccccttcgc ctcgtgcttg agttgaggcc
      421 tggcttgggc gctggggccg ccgcgtgcga atctggtggc accttcgcgc ctgtctcgct
      481 gctttcgata agtctctagc catttaaaat ttttgatgac ctgctgcgac gctttttttc
      541 tggcaagata gtcttgtaaa tgcgggccaa gatctgcaca ctggtatttc ggtttttggg
      601 gccgcgggcg gcgacggggc ccgtgcgtcc cagcgcacat gttcggcgag gcggggcctg
      661 cgagcgcggc caccgagaat cggacggggg tagtctcaag ctggccggcc tgctctggtg
      721 cctggcctcg cgccgccgtg tatcgccccg ccctgggcgg caaggctggc ccggtcggca
      781 ccagttgcgt gagcggaaag atggccgctt cccggccctg ctgcagggag ctcaaaatgg
      841 aggacgcggc gctcgggaga gcgggcgggt gagtcaccca cacaaaggaa aagggccttt
      901 ccgtcctcag ccgtcgcttc atgtgactcc acggagtacc gggcgccgtc caggcacctc
      961 gattagttct cgagcttttg gagtacgtcg tctttaggtt ggggggaggg gttttatgcg
     1021 atggagtttc cccacactga gtgggtggag actgaagtta ggccagcttg gcacttgatg
     1081 taattctcct tggaatttgc cctttttgag tttggatctt ggttcattct caagcctcag
     1141 acagtggttc aaagtttttt tcttccattt caggtgtcgt gacgctagcg ctaccggact
     1201 cagatctcga gctcaagctt cgaattcgcc accatggtgg gccggaacag cgctattgcc
     1261 gctggagtgt gcggcgccct gttcatcggc tactgcatct acttcgacag gaagagaagg
     1321 agcgacccca acttcaagaa caggctgagg gagagaagga agaagcagaa gctggccaag
     1381 gagagagccg gcctgagcaa gctgcccgac ctgaaggacg ccgaggccgt gcagaagttc
     1441 ggtaccgtga gcaagggcga ggagaataac atggccatca tcaaggagtt catgcgcttc
     1501 aaggtgcgca tggagggctc cgtgaacggc cacgagttcg agatcgaggg cgagggcgag
     1561 ggccgcccct acgagggctt tcagaccgtt aagctgaagg tgaccaaggg tggccccctg
     1621 cccttcgcct gggacatctt gtcccctcag ttcacctacg gctccaaggc ctacgtgaag
     1681 caccccgccg acatccccga ctacctcaag ctgtccttcc ccgagggctt caagtgggag
     1741 cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga ctcaggactc ctccctgcag
     1801 gacggcgagt tcatctacaa ggtgaagctg cgcggcacca acttcccctc cgacggcccc
     1861 gtaatgcaga agaagaccat gggcatggag gcctcctccg agcggatgta ccccgaggac
     1921 ggcgccctga agggcgagga caagctcagg ctgaagctga aggacggcgg ccactacacc
     1981 tccgaggtca agaccaccta caaggccaag aagcccgtgc agttgcccgg cgcctacatc
     2041 gtcgacatca agttggacat cacctcccac aacgaggact acaccatcgt ggaacagtac
     2101 gaacgcgccg agggccgcca ctccaccggc ggcatggacg agctgtacaa ggatatctcc
     2161 ggaggtatgg tcttcacact cgaagatttc gttggggact ggcgacagac agccggctac
     2221 aacctggacc aagtccttga acagggaggt gtgtccagtt tgtttcagaa tctcggggtg
     2281 tccgtaactc cgatccaaag gattgtcctg agcggtgaaa atgggctgaa gatcgacatc
     2341 catgtcatca tcccgtatga aggtctgagc ggcgaccaaa tgggccagat cgaaaaaatt
     2401 tttaaggtgg tgtaccctgt ggatgatcat cactttaagg tgatcctgca ctatggcaca
     2461 ctggtaatcg acggggttac gccgaacatg atcgactatt tcggacggcc gtatgaaggc
     2521 atcgccgtgt tcgacggcaa aaagatcact gtaacaggga ccctgtggaa cggcaacaaa
     2581 attatcgacg agcgcctgat caaccccgac ggctccctgc tgttccgagt aaccatcaac
     2641 ggagtgaccg gctggcggct gtgcgaacgc attctggcgt aaggatcccg cgactctaga
     2701 tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac ctcccacacc
     2761 tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag
     2821 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     2881 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttaaggc gtaaattgta
     2941 agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac
     3001 caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg
     3061 agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa
     3121 gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt
     3181 tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt
     3241 agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga
     3301 gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc
     3361 gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga
     3421 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     3481 ccctgataaa tgcttcaata atattgaaaa aggaagagtc ctgaggcgga aagaaccagc
     3541 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     3601 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     3661 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     3721 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     3781 taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt
     3841 agtgaggagg cttttttgga ggcctaggct tttgcaaaga tcgatcaaga gacaggatga
     3901 ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg
     3961 gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg
     4021 ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc
     4081 ctgaatgaac tgcaagacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct
     4141 tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa
     4201 gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg
     4261 gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa
     4321 gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat
     4381 gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg
     4441 agcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc
     4501 atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac
     4561 cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg
     4621 gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc
     4681 tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag
     4741 cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg
     4801 gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc
     4861 tggagttctt cgcccaccct agggggaggc taactgaaac acggaaggag acaataccgg
     4921 aaggaacccg cgctatgacg gcaataaaaa gacagaataa aacgcacggt gttgggtcgt
     4981 ttgttcataa acgcggggtt cggtcccagg gctggcactc tgtcgatacc ccaccgagac
     5041 cccattgggg ccaatacgcc cgcgtttctt ccttttcccc accccacccc ccaagttcgg
     5101 gtgaaggccc agggctcgca gccaacgtcg gggcggcagg ccctgccata gcctcaggtt
     5161 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     5221 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     5281 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     5341 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     5401 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     5461 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     5521 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     5581 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     5641 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     5701 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     5761 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     5821 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     5881 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     5941 tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc
     6001 gtattaccgc catgcattag ttattaatga gtaattcata caaaaggact cgcccctgcc
     6061 ttggggaatc ccagggaccg tcgttaaact cccactaacg tagaacccag agatcgctgc
     6121 gttcccgccc cctcacccgc ccgctctcgt catcactgag gtggagaaga gcatgcgtga
     6181 g
//
