LOCUS       sequence-345344-        4759 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  meGFP targeted to the mitochondrial matrix with a human COX8 
            presequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4759)
  TITLE     Fluorescent plasmids for live cell imaging in primary neurons
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4759)
  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: 
            "Unpublished"
COMMENT     Sequence Label: mito-meGFP
FEATURES             Location/Qualifiers
     source          1..4759
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..204
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     155..175
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             233..307
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     regulatory      335..344
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(341..343,344..346,347..1060)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(386..407)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(647..666)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     994..1015
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1183..1304
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1220..1239)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1274..1293
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1311..1766)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1448..1469)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1660..1679
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1793..1897
                     /gene="bla"
                     /label=AmpR promoter
     promoter        1899..2256
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1899..2090
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1922..1942)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2107..2242
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2291..3085
                     /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             2291..3085
                     /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(2345..2364)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2955..2974
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3273..3292)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3317..3364
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      3693..4281
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4182..4201
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        4456..4759
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
       61 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
      121 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
      181 tgggaggtct atataagcag agctggttta gtgaaccgtc agatccgcta gcatgtccgt
      241 cctgacgccg ctgctgctgc ggggcttgac aggctcggcc cggcggctcc cagtgccgcg
      301 cgccaagatc cattcgttgg gggatccacc ggtcgccacc atggtgagca agggcgagga
      361 gctgttcacc ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa
      421 gttcagcgtg tccggcgagg gcgagggcga tgccacctac ggcaagctga ccctgaagtt
      481 catctgcacc accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta
      541 cggcgtgcag tgcttcagcc gctaccccga ccacatgaag cagcacgact tcttcaagtc
      601 cgccatgccc gaaggctacg tccaggagcg caccatcttc ttcaaggacg acggcaacta
      661 caagacccgc gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa
      721 gggcatcgac ttcaaggagg acggcaacat cctggggcac aagctggagt acaactacaa
      781 cagccacaac gtctatatca tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa
      841 gatccgccac aacatcgagg acggcagcgt gcagctcgcc gaccactacc agcagaacac
      901 ccccatcggc gacggccccg tgctgctgcc cgacaaccac tacctgagca cccagtccaa
      961 gctgagcaaa gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc
     1021 cgccgggatc actctcggca tggacgagct gtacaagtaa agcggccgcg actctagatc
     1081 ataatcagcc ataccacatt tgtagaggtt ttacttgctt taaaaaacct cccacacctc
     1141 cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt tattgcagct
     1201 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     1261 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttaaggcgt aaattgtaag
     1321 cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca
     1381 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag
     1441 tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg
     1501 gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt
     1561 tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag
     1621 agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc
     1681 gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc
     1741 gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac
     1801 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     1861 ctgataaatg cttcaataat attgaaaaag gaagagtcct gaggcggaaa gaaccagctg
     1921 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     1981 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     2041 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     2101 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     2161 atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag
     2221 tgaggaggct tttttggagg cctaggcttt tgcaaagatc gatcaagaga caggatgagg
     2281 atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga
     2341 gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt
     2401 ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct
     2461 gaatgaactg caagacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg
     2521 cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt
     2581 gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc
     2641 tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc
     2701 gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga
     2761 tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgag
     2821 catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat
     2881 ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg
     2941 ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc
     3001 tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta
     3061 tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg
     3121 acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     3181 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     3241 gagttcttcg cccaccctag ggggaggcta actgaaacac ggaaggagac aataccggaa
     3301 ggaacccgcg ctatgacggc aataaaaaga cagaataaaa cgcacggtgt tgggtcgttt
     3361 gttcataaac gcggggttcg gtcccagggc tggcactctg tcgatacccc accgagaccc
     3421 cattggggcc aatacgcccg cgtttcttcc ttttccccac cccacccccc aagttcgggt
     3481 gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc ctcaggttac
     3541 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
     3601 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     3661 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     3721 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     3781 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
     3841 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     3901 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     3961 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     4021 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     4081 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     4141 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     4201 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     4261 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     4321 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     4381 attaccgcca tgcattagtt attaatagta atcaattacg gggtcattag ttcatagccc
     4441 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
     4501 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
     4561 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
     4621 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
     4681 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
     4741 agtcatcgct attaccatg
//
