LOCUS       sequence-245546-        4854 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses mitochondrial roGFP2 (mt-roGFP2) in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4854)
  AUTHORS   Ivashchenko O, Van Veldhoven PP, Brees C, Ho YS, Terlecky SR, 
            Fransen M
  TITLE     Intraperoxisomal redox balance in mammalian cells: oxidative stress 
            and interorganellar cross-talk.
  JOURNAL   Mol Biol Cell. 2011 May;22(9):1440-51. doi: 10.1091/mbc.E10-11-0919.
            Epub 2011 Mar 3.
  PUBMED    21372177
REFERENCE   2  (bases 1 to 4854)
  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: 
            "10.1091/mbc.E10-11-0919"; journalName: "Mol Biol Cell"; date: 
            "2011-05"; volume: "22"; issue: "9"; pages: "1440-51"
COMMENT     Sequence Label: pMF1762
FEATURES             Location/Qualifiers
     source          1..4854
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     10..30
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             88..162
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     CDS             184..255
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="SVLTPLLLRGLTGSARRLPVPRAK"
     CDS             join(292..294,295..297,298..1011)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFISTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNCHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTCSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(337..358)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(598..617)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     945..966
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1133..1254
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1170..1189)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1224..1243
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1261..1716)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1398..1419)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1610..1629
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1743..1847
                     /gene="bla"
                     /label=AmpR promoter
     promoter        1849..2206
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1849..2040
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1872..1892)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2057..2192
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2241..3035
                     /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             2241..3035
                     /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(2295..2314)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2905..2924
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3223..3242)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3267..3314
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      3643..4231
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4132..4151
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        4406..4709
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        join(4710..4854,1..59)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
ORIGIN
        1 ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagagctg
       61 gtttagtgaa ccgtcagatc cgctagcatg tccgtcctga cgccgctgct gctgcggggc
      121 ttgacaggct cggcccggcg gctcccagtg ccgcgcgcca agatccattc gttgggggat
      181 ctgtccgtcc tgacgccgct gctgctgcgg ggcttgacag gctcggcccg gcggctccca
      241 gtgccgcgcg ccaagatcca ttcgttgggg gatctcgagc tcaagcttac catggtgagc
      301 aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta
      361 aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg
      421 accctgaagt tcatctccac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc
      481 accctgacct acggcgtgca gtgcttcagc cgctaccctg accacatgaa gcagcacgac
      541 ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac
      601 gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc
      661 atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag
      721 tacaactaca actgccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag
      781 gtgaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac
      841 cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc
      901 acctgctccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag
      961 ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaagta agcggccgcg
     1021 actctagatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt taaaaaacct
     1081 cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt
     1141 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     1201 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttaaggcgt
     1261 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
     1321 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
     1381 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
     1441 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
     1501 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
     1561 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
     1621 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
     1681 cacccgccgc gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt cggggaaatg
     1741 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     1801 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtcct gaggcggaaa
     1861 gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg
     1921 cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg
     1981 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc
     2041 gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca
     2101 tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt
     2161 ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaagatc gatcaagaga
     2221 caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg
     2281 cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg
     2341 ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt
     2401 ccggtgccct gaatgaactg caagacgagg cagcgcggct atcgtggctg gccacgacgg
     2461 gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat
     2521 tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat
     2581 ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg
     2641 accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg
     2701 atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc
     2761 tcaaggcgag catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc
     2821 cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg
     2881 tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg
     2941 gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca
     3001 tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac
     3061 cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga
     3121 aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga
     3181 tctcatgctg gagttcttcg cccaccctag ggggaggcta actgaaacac ggaaggagac
     3241 aataccggaa ggaacccgcg ctatgacggc aataaaaaga cagaataaaa cgcacggtgt
     3301 tgggtcgttt gttcataaac gcggggttcg gtcccagggc tggcactctg tcgatacccc
     3361 accgagaccc cattggggcc aatacgcccg cgtttcttcc ttttccccac cccacccccc
     3421 aagttcgggt gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc
     3481 ctcaggttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     3541 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     3601 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     3661 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     3721 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     3781 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     3841 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     3901 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     3961 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     4021 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     4081 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     4141 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     4201 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     4261 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     4321 ggataaccgt attaccgcca tgcattagtt attaatagta atcaattacg gggtcattag
     4381 ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct
     4441 gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc
     4501 caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg
     4561 cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat
     4621 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
     4681 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc
     4741 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
     4801 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccg
//
