LOCUS       sequence-234984-        5850 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fluorescent vector with zebrafish pax3a enhancer 5; Tol2 version.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5850)
  AUTHORS   Chong-Morrison V, Mayes S, Simoes FC, Senanayake U, Carroll DS, 
            Riley PR, Wilson SW, Sauka-Spengler T
  TITLE     Ac/Ds transposition for CRISPR/dCas9-SID4x epigenome modulation in 
            zebrafish.
  JOURNAL   Biol Open. 2023 Jun 15;12(6):bio059995. doi: 10.1242/bio.059995. 
            Epub 2023 Jun 27.
  PUBMED    37367831
REFERENCE   2  (bases 1 to 5850)
  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: "Biol 
            Open."; date: "2023-06-15"; volume: "12(6):bio059995. doi"; pages: "
            10.1242/bio.059995. Epub 2023 Jun 27"
COMMENT     Sequence Label: pVC-Tol2-Pax3a_enh5-E1b:eGFP-Tol2
FEATURES             Location/Qualifiers
     source          1..5850
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    73..94
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(109..114,115..132,133..139)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    147..163
                     /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)."
     primer_bind     152..174
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     171..187
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     171..187
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     206..226
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        208..226
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     263..279
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     263..279
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     misc_feature    join(313..324,325..512)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     intron          1294..1866
                     /label=beta-globin intron
                     /note="intron from rabbit beta-globin gene"
     regulatory      1940..1949
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1946..1948,1949..1951,1952..2665)
                     /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             join(1946..1948,1949..1951,1952..2662)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1991..2012)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2252..2271)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2599..2620
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2875..2956
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2912..2931)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     misc_feature    join(3065..3202,3203..3214)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(3273..3290)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3273..3289)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3282..3304)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3431..3885
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3517..3536)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3727..3748
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3911..4015
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4016..4084,4085..4876)
                     /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     complement(4234..4253)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5047..5635
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5536..5555
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5789..5806
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga
       61 gcgcaacgca attaatgtga gttagctcac tcattaggca ccccaggctt tacactttat
      121 gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag
      181 ctatgaccat gattacgcca agcgcgcaat taaccctcac taaagggaac aaaagctgga
      241 gctccaccgc ggtggcggcc gctctagaac tagtggatct gctgggcttg ctgaaggtag
      301 ggggtcaaga accagaggtg taaagtactt gagtaatttt acttgattac tgtacttaag
      361 tattattttt ggggattttt actttacttg agtacaatta aaaatcaata cttttacttt
      421 tacttaatta cattttttta gaaaaaaaag tactttttac tccttacaat tttatttaca
      481 gtcaaaaagt acttattttt tggagatcac ttgggcccgg ctcgagccac acaattgtcc
      541 caacacaaat cgattaaatt aaaataatcg ttttattttt aatccacttc aatttgtaaa
      601 aacaattaat taaaaaagta gtttgaacaa gcagcaaaag taatattttc agtatacaaa
      661 gcgccaaaag cactgcagat aatttcttat ttgtttttgc gccctcttgc ggacagaaga
      721 atattgcagg gaaagccgct ttgagaggtg cgtgaatctt catgtctctc cggctttacc
      781 acctgcagag aggagcgcta ctggtagcca tgatcacctg agagcaataa tgccggagct
      841 caggccccgg ggccaaggtt caccgcgggg gcaataaact cctgtccccc tgccatcatg
      901 taaagacatt accgcgccct gcggccctca caacaaacac tccatttcca aaggctctct
      961 gccgcttact gggtcgcttt atttaccctg tgtcatgcaa aagtcttaat ttcctttgaa
     1021 tcccagtgct gctgtattgc cttgagttac ttacattcat taccaccgtt taattagtac
     1081 catcaaacgc ttcacggttt acagtggctc aatgtgacat tagtcgagac tgttaaactc
     1141 attagcaaat actgataagg gggtaaaggt gagatggctg ctagcaagct tgcggcgata
     1201 tcgactctag agggtatata atggatccca tcgcgtctca gcctcacttt gagctcctcc
     1261 acacgaattt cgaccgatcc tgagaacttc agggtgagtt tggggaccct tgattgttct
     1321 ttctttttcg ctattgtaaa attcatgtta tatggagggg gcaaagtttt cagggtgttg
     1381 tttagaatgg gaagatgtcc cttgtatcac catggaccct catgataatt ttgtttcttt
     1441 cactttctac tctgttgaca accattgtct cctcttattt tcttttcatt ttctgtaact
     1501 ttttcgttaa actttagctt gcatttgtaa cgaattttta aattcacttt tgtttatttg
     1561 tcagattgta agtactttct ctaatcactt ttttttcaag gcaatcaggg tatattatat
     1621 tgtacttcag cacagtttta gagaacaatt gttataatta aatgataagg tagaatattt
     1681 ctgcatataa attctggctg gcgtggaaat attcttattg gtagaaacaa ctacaccctg
     1741 gtcatcatcc tgcctttctc tttatggtta caatgatata cactgtttga gatgaggata
     1801 aaatactctg agtccaaacc gggcccctct gctaaccatg ttcatgcctt cttctctttc
     1861 ctacagctcc tgggcaacgt gctggttgtt gtgctgtctc atcattttgg caaagaattc
     1921 ctcgacggat ccaccggtcg ccaccatggt gagcaagggc gaggagctgt tcaccggggt
     1981 ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg
     2041 cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcatct gcaccaccgg
     2101 caagctgccc gtgccctggc ccaccctcgt gaccaccctg acctacggcg tgcagtgctt
     2161 cagccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg
     2221 ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga
     2281 ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa
     2341 ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta
     2401 tatcatggcc gacaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacat
     2461 cgaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg
     2521 ccccgtgctg ctgcccgaca accactacct gagcacccag tccgccctga gcaaagaccc
     2581 caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg ggatcactct
     2641 cggcatggac gagctgtaca agtaaagcgg ccgcgactct agaattccaa ctgagcgccg
     2701 gtcgctacca ttaccaactt gtctggtgtc aaaaataata ggcctactag taccggatct
     2761 agataactga tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac
     2821 ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg
     2881 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     2941 gcattttttt cactgccaat tgcattcatt ttatgtttcg ggttcagggg gaggtgtggg
     3001 aggtttttta aagcaagtaa aacctctaca aatgtggtac ccgggtttaa acgcatgcag
     3061 atctaatact caagtacaat tttaatggag tactttttta cttttactca agtaagattc
     3121 tagccagata cttttacttt taattgagta aaattttccc taagtacttg tactttcact
     3181 tgagtaaaat ttttgagtac tttttacacc tctgtcaaga accatatgcc ggtacccaat
     3241 tcgccctata ggacgtcgta ttacgcgcgc tcactggccg tcgttttaca acgtcgtgac
     3301 tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc
     3361 tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat
     3421 ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     3481 agcgtgacca ccacacccgc cgcgccctag cgcccgctcc tttcgctttc ttcccttcct
     3541 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     3601 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac
     3661 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     3721 ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt
     3781 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     3841 aaaaatttaa cgcgaatttt aacaaaatat taacgcttac aatttaggtg gcacttttcg
     3901 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     3961 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
     4021 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     4081 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     4141 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     4201 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     4261 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     4321 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     4381 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     4441 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     4501 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     4561 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     4621 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     4681 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     4741 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     4801 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     4861 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     4921 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     4981 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     5041 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     5101 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     5161 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     5221 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     5281 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     5341 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     5401 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     5461 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     5521 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     5581 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     5641 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     5701 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
     5761 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg
     5821 cccaatacgc aaaccgcctc tccccgcgcg
//
