LOCUS       sequence-161603-        4916 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of redox sensitive GFP with a mitochondrial targeting 
            sequence in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4916)
  AUTHORS   Hanson GT, Aggeler R, Oglesbee D, Cannon M, Capaldi RA, Tsien RY, 
            Remington SJ
  TITLE     Investigating mitochondrial redox potential with redox-sensitive 
            green fluorescent protein indicators.
  JOURNAL   J Biol Chem. 2004 Mar 26;279(13):13044-53. Epub 2004 Jan 13.
  PUBMED    14722062
REFERENCE   2  (bases 1 to 4916)
  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 Biol 
            Chem."; date: "2004-03-26"; volume: "279(13)"; pages: "13044-53. 
            Epub 2004 Jan 13"
COMMENT     Sequence Label: pEGFP-N1-mt-ro1GFP
FEATURES             Location/Qualifiers
     source          1..4916
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..441
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        442..645
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     596..616
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      933..942
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(939..941,942..944,945..1658)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFISTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNCHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTCSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(984..1005)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1245..1264)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1592..1613
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1781..1902
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1818..1837)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1872..1891
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1909..2364)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2046..2067)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2258..2277
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2391..2495
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2497..2854
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2497..2688
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2520..2540)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2705..2840
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2889..3683
                     /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             2889..3683
                     /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(2943..2962)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3553..3572
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3871..3890)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3915..3962
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4291..4879
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4780..4799
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc
       61 gtattaccgc catgcattag ttattaatag taatcaatta cggggtcatt agttcatagc
      121 ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc
      181 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg
      241 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat
      301 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc
      361 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta
      421 ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag
      481 cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt
      541 tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa
      601 atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt
      661 cagatccgct agcgctaccg gactcagatc tcgagctcga attcgggctg aaggagactt
      721 gggggcaccc gcgtcgtgcc tcctgggttg tgaggagtcg ccgctgccgc cactgcctgt
      781 gcttcatgag gaagatgctc gccgccgtct cccgcgtgct gtctggcgct tctcagaagc
      841 cggcaagcag agtgctggta gcatcccgta attttgcaaa tgatgctaca tttctgcagt
      901 cgacggtacc gcgggcccgg gatccaccgg tcgccaccat ggtgagcaag ggcgaggagc
      961 tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt
     1021 tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca
     1081 tctccaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgtcctacg
     1141 gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg
     1201 ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca
     1261 agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg
     1321 gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaact
     1381 gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg aacttcaaga
     1441 tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc
     1501 ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc tgctccgccc
     1561 tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg
     1621 ccgggatcac tctcggcatg gacgagctgt acaagtaaag cggccgcgac tctagatcat
     1681 aatcagccat accacatttg tagaggtttt acttgcttta aaaaacctcc cacacctccc
     1741 cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt aacttgttta ttgcagctta
     1801 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     1861 gcattctagt tgtggtttgt ccaaactcat caatgtatct taaggcgtaa attgtaagcg
     1921 ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat
     1981 aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg
     2041 ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc
     2101 gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt
     2161 tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag
     2221 cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg
     2281 gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc
     2341 ttaatgcgcc gctacagggc gcgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     2401 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     2461 gataaatgct tcaataatat tgaaaaagga agagtcctga ggcggaaaga accagctgtg
     2521 gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     2581 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
     2641 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
     2701 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     2761 tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg
     2821 aggaggcttt tttggaggcc taggcttttg caaagatcga tcaagagaca ggatgaggat
     2881 cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga
     2941 ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc
     3001 ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga
     3061 atgaactgca agacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg
     3121 cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc
     3181 cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg
     3241 atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga
     3301 aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc
     3361 tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgagca
     3421 tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg
     3481 tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct
     3541 atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg
     3601 accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc
     3661 gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac
     3721 gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
     3781 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
     3841 gttcttcgcc caccctaggg ggaggctaac tgaaacacgg aaggagacaa taccggaagg
     3901 aacccgcgct atgacggcaa taaaaagaca gaataaaacg cacggtgttg ggtcgtttgt
     3961 tcataaacgc ggggttcggt cccagggctg gcactctgtc gataccccac cgagacccca
     4021 ttggggccaa tacgcccgcg tttcttcctt ttccccaccc caccccccaa gttcgggtga
     4081 aggcccaggg ctcgcagcca acgtcggggc ggcaggccct gccatagcct caggttactc
     4141 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     4201 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     4261 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     4321 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     4381 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     4441 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     4501 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     4561 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     4621 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     4681 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     4741 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     4801 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     4861 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggcct
//
