LOCUS       sequence-14313-e        4925 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mitochondrial localization of EGFP, for real-time imaging of 
            mitochondria in transgenic zebrafish.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4925)
  AUTHORS   Kim MJ, Kang KH, Kim CH, Choi SY
  TITLE     Real-time imaging of mitochondria in transgenic zebrafish expressing
            mitochondrially targeted GFP.
  JOURNAL   Biotechniques. 2008 Sep . 45(3):331-4.
  PUBMED    18778258
REFERENCE   2  (bases 1 to 4925)
  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: 
            "Biotechniques. 2008 Sep . 45(3):331-4."
COMMENT     Sequence Label: pSCAC-69
FEATURES             Location/Qualifiers
     source          1..4925
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(34..462)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(144..165)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     356..375
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     589..611
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     603..620
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     604..620
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     630..649
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        630..648
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    join(679..690,691..828)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     polyA_signal    950..1084
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(987..1006)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1041..1060
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(1129..1842,1843..1845,1846..1848))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(1132..1842,1843..1845,1846..1848))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1174..1195)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1523..1542
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1782..1803
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      1845..1854
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     misc_feature    join(2455..2642,2643..2654)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(2688..2704)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2688..2704)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(2741..2761)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2741..2759)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2780..2796)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2780..2796)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2793..2815)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2804..2820
                     /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(2828..2834,2835..2852,2853..2858))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2873..2894
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3011..3028)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3182..3770)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3262..3281)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3941..4732,4733..4801))
                     /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     4564..4583
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4802..4906)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ccacctaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
       61 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
      121 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
      181 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
      241 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
      301 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga
      361 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
      421 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcccattcg ccattcaggc
      481 tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga
      541 aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac
      601 gttgtaaaac gacggccagt gagcgcgcgt aatacgactc actatagggc gaattgggta
      661 ccggcatatg gttcttgaca gaggtgtaaa aagtactcaa aaattttact caagtgaaag
      721 tacaagtact tagggaaaat tttactcaat taaaagtaaa agtatctggc tagaatctta
      781 cttgagtaaa agtaaaaaag tactccatta aaattgtact tgagtattag atctgatcta
      841 gaggatcata atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc
      901 acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat
      961 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     1021 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg
     1081 gatcatcatc gatgaattaa ttcgagctcc accgcggtgg cggccgcttt acttgtacag
     1141 ctcgtccatg ccgagagtga tcccggcggc ggtcacgaac tccagcagga ccatgtgatc
     1201 gcgcttctcg ttggggtctt tgctcagggc ggactgggtg ctcaggtagt ggttgtcggg
     1261 cagcagcacg gggccgtcgc cgatgggggt gttctgctgg tagtggtcgg cgagctgcac
     1321 gctgccgtcc tcgatgttgt ggcggatctt gaagttcacc ttgatgccgt tcttctgctt
     1381 gtcggccatg atatagacgt tgtggctgtt gtagttgtac tccagcttgt gccccaggat
     1441 gttgccgtcc tccttgaagt cgatgccctt cagctcgatg cggttcacca gggtgtcgcc
     1501 ctcgaacttc acctcggcgc gggtcttgta gttgccgtcg tccttgaaga agatggtgcg
     1561 ctcctggacg tagccttcgg gcatggcgga cttgaagaag tcgtgctgct tcatgtggtc
     1621 ggggtagcgg ctgaagcact gcacgccgta ggtcagggtg gtcacgaggg tgggccaggg
     1681 cacgggcagc ttgccggtgg tgcagatgaa cttcagggtc agcttgccgt aggtggcatc
     1741 gccctcgccc tcgccggaca cgctgaactt gtggccgttt acgtcgccgt ccagctcgac
     1801 caggatgggc accaccccgg tgaacagctc ctcgcccttg ctcaccatgg tggcgaccgg
     1861 tcgcgtaacg aggttggctc gctgggtgat cgtggatccc cgcagaaccg gagcggcgcg
     1921 gacgcgagct agtcccctca gaagtccaga catggtaagc ttcagctaga actcgccgca
     1981 gacccggtga ggaagagagc gaaccgggcc ttaaccaccc tttatatagc cgcctctact
     2041 gggcggggat taaccatgac atcatcaagt ccagaattcc caaagtgcta gaggcggggt
     2101 cttgacagaa cattcagcct gtaagcgtct tgtatactac aactcccagt agcactaggg
     2161 cggatgcagc tgttgagtga aacgcgcgag ttatcacgaa gttagggcga aaggaagggt
     2221 ggcactccct agtgcgtcat aagctagctt gcatgcctga gaatttcaga atgtaatgat
     2281 acctttgtta gataaggctg tttacatctg atagtggacc ttaagccgac acttaaatga
     2341 taaaaacggc aaagaattgc aagtttgatt tgcattggaa agggtcgctg gcttttgtgt
     2401 tacacgcccc ttatttgtgc ttgattagat gatccccctg ctcgagccgg gcccaagtga
     2461 tctccaaaaa ataagtactt tttgactgta aataaaattg taaggagtaa aaagtacttt
     2521 tttttctaaa aaaatgtaat taagtaaaag taaaagtatt gatttttaat tgtactcaag
     2581 taaagtaaaa atccccaaaa ataatactta agtacagtaa tcaagtaaaa ttactcaagt
     2641 actttacacc tctggttctt gaccccctac cttcagcaag cccagcagat ccactagttc
     2701 tagagcggcc gccaccgcgg tggagctcca gcttttgttc cctttagtga gggttaattg
     2761 cgcgcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     2821 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     2881 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     2941 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     3001 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     3061 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     3121 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     3181 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     3241 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     3301 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     3361 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     3421 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     3481 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     3541 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     3601 ctaactacgg ctacactaga aggacagtat ttggtatctg cgctctgctg aagccagtta
     3661 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     3721 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     3781 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     3841 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     3901 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     3961 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     4021 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     4081 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     4141 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     4201 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     4261 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     4321 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     4381 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     4441 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     4501 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     4561 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     4621 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     4681 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     4741 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     4801 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     4861 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     4921 aagtg
//
