LOCUS       sequence-329887-        5685 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of mito-roGFP2 (mitochondrial matrix targeted roGFP2) 
            used for monitoring dynamcs of mitochondrial Redox potential.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5685)
  AUTHORS   Lewis TL Jr, Kwon SK, Lee A, Shaw R, Polleux F
  TITLE     MFF-dependent mitochondrial fission regulates presynaptic release 
            and axon branching by limiting axonal mitochondria size.
  JOURNAL   Nat Commun. 2018 Nov 27;9(1):5008. doi: 10.1038/s41467-018-07416-2.
  PUBMED    30479337
REFERENCE   2  (bases 1 to 5685)
  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: "Nat 
            Commun."; date: "2018-11-27"; volume: "9(1):5008. doi"; pages: " 
            10.1038/s41467-018-07416-2"
COMMENT     Sequence Label: pCAG roGFP2-mito
FEATURES             Location/Qualifiers
     source          1..5685
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        34..413
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        415..692
                     /label=chicken beta-actin promoter
     intron          693..1709
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1633..1655
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1717..1736
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1770..1844
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     CDS             join(1872..1874,1875..2585)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FISTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNCHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTCSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1914..1935)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2175..2194)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2522..2543
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2706..2725)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2771..2826
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2825..2844)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3217..3233)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3217..3233)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3230..3252)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3241..3257
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(3265..3271,3272..3289,3290..3295))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3310..3331
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3390..3585
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3436..3571
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3591..3725
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3628..3647)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3682..3701
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3793..3810)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3964..4552)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4044..4063)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4723..5514,5515..5583))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5346..5365
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(5584..5685,1..3))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gcgcacattt ccccgaaaag tgccacctgg gtcgacattg attattgact agttattaat
       61 agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac
      121 ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa
      181 tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt
      241 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc
      301 ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat
      361 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtcgagg
      421 tgagccccac gttctgcttc actctcccca tctccccccc ctccccaccc ccaattttgt
      481 atttatttat tttttaatta ttttgtgcag cgatgggggc gggggggggg ggggggcgcg
      541 cgccrggsgg gggggggsgg ggsggggggs ggggsggggs grggcggaga ggtgcggcgg
      601 cagccaatca gagcggcgcg ctccgaaagt ttccttttat ggcgaggcgg cggcggcggc
      661 ggccctataa aaagcgaagc gcgcggcggg cgggagtcgc tgcgcgctgc cttcgccccg
      721 tgccccgctc cgccgccgcc tcgcgccgcc cgccccggct ctgactgacc gcgttactcc
      781 cacaggtgag cgggcgggac ggcccttctc ctccgggctg taattagcgc ttggtttaat
      841 gacggcttgt ttcttttctg tggctgcgtg aaagccttga ggggctccgg gagggccctt
      901 tgtgcggggg gagcggctcg gggggtgcgt gcgtgtgtgt gtgcgtgggg agcgccgcgt
      961 gcggctccgc gctgcccggc ggctgtgagc gctgcgggcg cggcgcgggg ctttgtgcgc
     1021 tccgcagtgt gcgcgagggg agcgcggccg ggggcggtgc cccgcggtgc ggggggggct
     1081 gcgaggggaa caaaggctgc gtgcggggtg tgtgcgtggg ggggtgagca gggggtgtgg
     1141 gcgcgtcggt cgggctgcaa ccccccctgc acccccctcc ccgagttgct gagcacggcc
     1201 cggcttcggg tgcggggctc cgtacggggc gtggcgcggg gctcgccgtg ccgggcgggg
     1261 ggtggcggca ggtgggggtg ccgggcgggg cggggccgcc tcgggccggg gagggctcgg
     1321 gggaggggcg cggcggcccc cggagcgccg gcggctgtcg aggcgcggcg agccgcagcc
     1381 attgcctttt atggtaatcg tgcgagaggg cgcagggact tcctttgtcc caaatctgtg
     1441 cggagccgaa atctgggagg cgccgccgca ccccctctag cgggcgcggg gcgaagcggt
     1501 gcggcgccgg caggaaggaa atgggcgggg agggccttcg tgcgtcgccg cgccgccgtc
     1561 cccttctccc tctccagcct cggggctgtc cgcgggggga cggctgcctt cgggggggac
     1621 ggggcagggc ggggttcggc ttctggcgtg tgaccggcgg ctctagagcc tctgctaacc
     1681 atgttcatgc cttcttcttt ttcctacagc tcctgggcaa cgtgctggtt attgtgctgt
     1741 ctcatcattt tggcaaagaa ttgctcgaga tgtccgtcct gacgccgctg ctgctgcggg
     1801 gcttgacagg ctcggcccgg cggctcccag tgccgcgcgc caagatccat tcgttggggg
     1861 atccaccggt agtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
     1921 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg
     1981 ccacctacgg caagctgacc ctgaagttca tctccaccac cggcaagctg cccgtgccct
     2041 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc
     2101 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
     2161 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
     2221 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     2281 tggggcacaa gctggagtac aactacaact gccacaacgt ctatatcatg gccgacaagc
     2341 agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc
     2401 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
     2461 acaaccacta cctgagcacc tgctccgccc tgagcaaaga ccccaacgag aagcgcgatc
     2521 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt
     2581 acaagactca cggtatggac gaattgtaca agtaagcggc cgcaattcac tcctcaggtg
     2641 caggctgcct atcagaaggt ggtggctggt gtggccaatg ccctggctca caaataccac
     2701 tgagatcttt ttccctctgc caaaaattat ggggacatca tgaagcccct tgagcatctg
     2761 acttctggct aataaaggaa atttattttc attgcaatag tgtgttggaa ttttttgtgt
     2821 ctctcactcg gaaggacata tgggagggca aatcatttaa aacatcagaa tgagtatttg
     2881 gtttagagtt tggcaacata tgcccatatg ctggctgcca tgaacaaagg ttggctataa
     2941 agaggtcatc agtatatgaa acagccccct gctgtccatt ccttattcca tagaaaagcc
     3001 ttgacttgag gttagatttt ttttatattt tgttttgtgt tatttttttc tttaacatcc
     3061 ctaaaatttt ccttacatgt tttactagcc agatttttcc tcctctcctg actactccca
     3121 gtcatagctg tccctcttct cttatggaga tccctcgacc tgcaccgtcg accagctggt
     3181 cgacggtgca ccgtcgacca gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     3241 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     3301 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     3361 gtcgggaaac ctgtcgtgcc agcggatccg catctcaatt agtcagcaac catagtcccg
     3421 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
     3481 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc
     3541 cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct aacttgttta
     3601 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     3661 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     3721 ggatccgctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     3781 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     3841 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     3901 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     3961 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     4021 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     4081 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     4141 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     4201 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     4261 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     4321 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     4381 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     4441 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     4501 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     4561 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     4621 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     4681 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     4741 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     4801 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     4861 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     4921 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     4981 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     5041 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     5101 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     5161 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     5221 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     5281 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     5341 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     5401 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     5461 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     5521 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     5581 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     5641 atacatattt gaatgtattt agaaaaataa acaaataggg gttcc
//
