LOCUS       sequence-456523-        4759 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Localization: Mitochondria, Excitation: 513, Emission: 527.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4759)
  AUTHORS   Rizzo MA, Davidson MW, Piston DW
  TITLE     Fluorescent protein tracking and detection: fluorescent protein 
            structure and color variants.
  JOURNAL   Cold Spring Harb Protoc. 2009 Dec;2009(12):pdb.top63. doi: 
            10.1101/pdb.top63.
  PUBMED    20150100
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: "Cold Spring
            Harb Protoc."; date: "2009-12"; volume: "2009(12):pdb.top63. doi"; 
            pages: " 10.1101/pdb.top63"
COMMENT     Sequence Label: EYFP-Mito-7
FEATURES             Location/Qualifiers
     source          1..4759
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     17..36
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        291..594
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        595..798
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     749..769
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             827..901
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     regulatory      929..938
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(935..937,938..940,941..1654)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(980..1001)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1241..1260)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1588..1609
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1777..1898
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1814..1833)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1868..1887
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1905..2360)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2042..2063)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2254..2273
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2387..2491
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2493..2850
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2493..2684
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2516..2536)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2701..2836
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2885..3679
                     /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             2885..3679
                     /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(2939..2958)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3549..3568
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3867..3886)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3911..3958
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      join(4287..4759,1..116)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
       61 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
      121 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
      181 ttcctgcgtt atcccctgat tctgtggata accgtattac cgccatgcat tagttattaa
      241 tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg cgttacataa
      301 cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata
      361 atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag
      421 tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc
      481 cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta
      541 tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg
      601 cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt
      661 ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca
      721 aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag
      781 gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcatgt ccgtcctgac
      841 gccgctgctg ctgcggggct tgacaggctc ggcccggcgg ctcccagtgc cgcgcgccaa
      901 gatccattcg ttgggggatc caccggtcgc caccatggtg agcaagggcg aggagctgtt
      961 caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag
     1021 cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg
     1081 caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccttcg gctacggcct
     1141 gcagtgcttc gcccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat
     1201 gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac
     1261 ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat
     1321 cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca
     1381 caacgtctat atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg
     1441 ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat
     1501 cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agctaccagt ccgccctgag
     1561 caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg
     1621 gatcactctc ggcatggacg agctgtacaa gtaaagcggc cgcgactcta gatcataatc
     1681 agccatacca catttgtaga ggttttactt gctttaaaaa acctcccaca cctccccctg
     1741 aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc agcttataat
     1801 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     1861 tctagttgtg gtttgtccaa actcatcaat gtatcttaag gcgtaaattg taagcgttaa
     1921 tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta accaataggc
     1981 cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt tgagtgttgt
     2041 tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca aagggcgaaa
     2101 aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa gttttttggg
     2161 gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat ttagagcttg
     2221 acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc
     2281 tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa
     2341 tgcgccgcta cagggcgcgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     2401 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     2461 aatgcttcaa taatattgaa aaaggaagag tcctgaggcg gaaagaacca gctgtggaat
     2521 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     2581 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     2641 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     2701 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     2761 tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga
     2821 ggcttttttg gaggcctagg cttttgcaaa gatcgatcaa gagacaggat gaggatcgtt
     2881 tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct
     2941 attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct
     3001 gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga
     3061 actgcaagac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc
     3121 tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg
     3181 gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc
     3241 aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca
     3301 tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga
     3361 cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg cgagcatgcc
     3421 cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga
     3481 aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca
     3541 ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg
     3601 cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct
     3661 tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc
     3721 aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga
     3781 atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc
     3841 ttcgcccacc ctagggggag gctaactgaa acacggaagg agacaatacc ggaaggaacc
     3901 cgcgctatga cggcaataaa aagacagaat aaaacgcacg gtgttgggtc gtttgttcat
     3961 aaacgcgggg ttcggtccca gggctggcac tctgtcgata ccccaccgag accccattgg
     4021 ggccaatacg cccgcgtttc ttccttttcc ccaccccacc ccccaagttc gggtgaaggc
     4081 ccagggctcg cagccaacgt cggggcggca ggccctgcca tagcctcagg ttactcatat
     4141 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     4201 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     4261 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     4321 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     4381 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     4441 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     4501 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     4561 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     4621 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     4681 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     4741 gtcggaacag gagagcgca
//
