LOCUS       sequence-115864-        5658 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Mito-CNL in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5658)
  AUTHORS   Takai A, Nakano M, Saito K, Haruno R, Watanabe TM, Ohyanagi T, Jin 
            T, Okada Y, Nagai T
  TITLE     Expanded palette of Nano-lanterns for real-time multicolor 
            luminescence imaging.
  JOURNAL   Proc Natl Acad Sci U S A. 2015 Apr 7;112(14):4352-6. doi: 
            10.1073/pnas.1418468112. Epub 2015 Mar 23.
  PUBMED    25831507
REFERENCE   2  (bases 1 to 5658)
  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: 
            "10.1073/pnas.1418468112"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2015-04-7- 7"; volume: "112"; issue: "14"; pages: "4352-6"
COMMENT     Sequence Label: pCNL-N1_Mito
FEATURES             Location/Qualifiers
     source          1..5658
                     /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"
     CDS             597..671
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     regulatory      699..708
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(705..707,708..710,711..1223)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYFSDNVYITADKQKNGIK
                     ANFKIRHNIE"
     primer_bind     complement(750..771)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1011..1030)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1358..1379
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2446..2567
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2483..2502)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2537..2556
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2574..3029)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2711..2732)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2923..2942
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3056..3160
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3162..3519
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3162..3353
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3185..3205)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3370..3505
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3554..4348
                     /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             3554..4348
                     /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(3608..3627)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4218..4237
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4536..4555)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4580..4627
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4956..5544
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5445..5464
                     /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 gctagcatgt
      601 ccgtcctgac gccgctgctg ctgcggggct tgacaggctc ggcccggcgg ctcccagtgc
      661 cgcgcgccaa gatccattcg ttgggggatc caccggtcgc caccatggtg agcaagggcg
      721 aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc
      781 acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga
      841 agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccctgt
      901 cctggggcgt gcagtgcttc gcccgctacc ccgaccacat gaagcagcac gacttcttca
      961 agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca
     1021 actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc
     1081 tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact
     1141 actttagcga caacgtctat atcaccgccg acaagcagaa gaacggcatc aaggccaact
     1201 tcaagatccg ccacaacatc gaggacggcg gcgtgcagct cgccgaccac taccagcaga
     1261 acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcacccagt
     1321 ccaagctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga
     1381 ccgccgccgg gggtaccaag gtgtacgacc ccgagcagag gaagaggatg atcaccggcc
     1441 cccagtggtg ggccaggtgc aagcagatga acgtgctgga cagcttcatc aactactacg
     1501 acagcgagaa gcacgccgag aacgccgtga tcttcctgca cggcaacgcc actagcagct
     1561 acctgtggag gcacgtggtg ccccacatcg agcccgtggc caggtgcatc atccccgatc
     1621 tgatcggcat gggcaagagc ggcaagagcg gcaacggcag ctacaggctg ctggaccact
     1681 acaagtacct gaccgcctgg ttcgagcttc tgaacctgcc caagaagatc atcttcgtgg
     1741 gccacgactg gggcgccgcc ctggccttcc actacgccta cgagcaccag gacaggatca
     1801 aggccatcgt gcacatggag agcgtggtgg acgtgatcga gagctgggac gagtggccag
     1861 acatcgagga ggacatcgcc ctgatcaaga gcgaggaggg cgagaagatg gtgctggaga
     1921 acaacttctt cgtggagacc gtgctgccca gcaagatcat gagaaagctg gagcccgagg
     1981 agttcgccgc ctacctggag cccttcaagg agaagggcga ggtgagaaga cccaccctga
     2041 gctggcccag agagatcccc ctggtgaagg gcggcaagcc cgacgtggtg cagatcgtga
     2101 gaaactacaa cgcctacctg agagccagcg acgacctgcc caagctgttc atcgagggcg
     2161 accccggctt cttcagcaac gccatcgtgg agggcgccaa gaagttcccc aacaccgagt
     2221 tcgtgaaggt gaagggcctg cacttcctcc aggaggacgc ccccgacgag atgggcaagt
     2281 acatcaagag cttcgtggag agagtgctga agaacgagca gtaggcggcc gcgactctag
     2341 atcataatca gccataccac atttgtagag gttttacttg ctttaaaaaa cctcccacac
     2401 ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt gtttattgca
     2461 gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt
     2521 tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttaagg cgtaaattgt
     2581 aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa
     2641 ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt
     2701 gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa
     2761 agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag
     2821 ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt
     2881 tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg
     2941 agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc
     3001 cgcgcttaat gcgccgctac agggcgcgtc aggtggcact tttcggggaa atgtgcgcgg
     3061 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     3121 accctgataa atgcttcaat aatattgaaa aaggaagagt cctgaggcgg aaagaaccag
     3181 ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt
     3241 atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca
     3301 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta
     3361 actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga
     3421 ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag
     3481 tagtgaggag gcttttttgg aggcctaggc ttttgcaaag atcgatcaag agacaggatg
     3541 aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt
     3601 ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt
     3661 gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc
     3721 cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc
     3781 ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga
     3841 agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat
     3901 ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca
     3961 agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga
     4021 tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc
     4081 gagcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat
     4141 catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga
     4201 ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg
     4261 ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt
     4321 ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa
     4381 gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg
     4441 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     4501 ctggagttct tcgcccaccc tagggggagg ctaactgaaa cacggaagga gacaataccg
     4561 gaaggaaccc gcgctatgac ggcaataaaa agacagaata aaacgcacgg tgttgggtcg
     4621 tttgttcata aacgcggggt tcggtcccag ggctggcact ctgtcgatac cccaccgaga
     4681 ccccattggg gccaatacgc ccgcgtttct tccttttccc caccccaccc cccaagttcg
     4741 ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag gccctgccat agcctcaggt
     4801 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     4861 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     4921 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     4981 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     5041 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     5101 gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     5161 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     5221 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     5281 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     5341 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     5401 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     5461 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     5521 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     5581 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     5641 cgtattaccg ccatgcat
//
